Synthesis of isofagomine and a new C6 pyrrolidine azasugar with potential biological activity

Article abstract of DOI:10.1055/s-2008-1078280

An efficient asymmetric synthesis of isofagomine, based on a precursor containing three differentiated hydroxyl functions, is described. The side product in the key alkylation step is converted into (2S,3R,4R)-2,4- bis(hydroxymethyl)-3-hydroxypyrrolidine, a new C₆ pyrrolidine azasugar, which inhibits α-glucosidase from yeast. Georg Thieme Verlag Stuttgart.

Full text of DOI:10.1055/s-2008-1078280

LETTER
2321
Synthesis of Isofagomine and a New C6 Pyrrolidine Azasugar with Potential
Biological Activity
S
ynthesis of Is
i
ofagom
e
ine and
a
N
tewC6
e
Pyrrolidin
r
A
zasugar E. R. Espeel,a Kathleen Piens,b Nico Callewaert,b Johan Van der Eycken*a
a
Laboratory for Organic and Bioorganic Synthesis, Department of Organic Chemistry, Ghent University, Krijgslaan 281 (S.4), 9000 Gent,
Belgium
b
Laboratory for Protein Biochemistry and Biomolecular Engineering, Department of Biochemistry, Physiology and Microbiology, Ghent
University, K.L. Ledeganckstraat 35, 9000 Gent, Belgium
Fax +32(9)2644998; E-mail: Johan.VanderEycken@UGent.be
Received 17 April 2008
HO
HO
Abstract: An efficient asymmetric synthesis of isofagomine, based
on a precursor containing three differentiated hydroxyl functions, is
described. The side product in the key alkylation step is converted
into (2S,3R,4R)-2,4-bis(hydroxymethyl)-3-hydroxypyrrolidine, a
new C6 pyrrolidine azasugar, which inhibits a-glucosidase from
yeast.
O
RCM
N
N
Boc
Boc
OH
rac-4
5
rac-6
OH
HO
O
Keywords: asymmetric synthesis, azasugars, glycosidase inhibi-
OH
tors, hydroboration, ring-closing metathesis
N
N
Boc
Boc
(S)-4
(S)-6
7
Isofagomine (1) is the most representative member of the
gluco-configured 1-azasugars: the ring oxygen, the ano-
meric carbon atom and the C-2 hydroxyl group in D-glu-
cose (2) are replaced by a methylene group, a nitrogen and
Scheme 1
A recently published synthesis4 of isofagomine (1) is
a hydrogen atom, respectively (Figure 1). Due to these based on N-Boc-5-hydroxy-3,4-didehydropiperidine
structural features, the stable protonated form of isofago- (rac-6) as a key intermediate (Scheme 1). It is obtained by
mine (1) shows obvious stereoelectronic similarity to the nucleophilic opening of the epoxide in vinyloxirane (rac-
oxocarbenium ion 3, which develops in the transition state 4) with allylamine, followed by Boc protection and ring-
during enzymatic hydrolysis of a glucoside while being closing metathesis (RCM). The didehydropiperidine rac-
stabilized by the enzyme.1,2 Therefore, isofagomine (1) is 6 is then resolved via kinetic enzymatic resolution and
subsequently used for the synthesis of various azasugars.4
a very potent reversible inhibitor of b-glucosidase.3
We modified this reaction sequence in such a way that ra-
cemic vinyloxirane (rac-4) was replaced with (S)-vinyl-
oxirane [(S)-4], thus rendering enzymatic resolution
obsolete. Enantiopure (S)-vinyloxirane [(S)-4]5 was
obtained via hydrolytic kinetic resolution with a chiral
(salen) CoIII complex.6 Using racemic rac-6 as a refer-
ence, an enantiomeric excess of 97% ee was determined
for (S)-6 by chiral HPLC analysis.7 The ee was improved
to ≥ 99% after recrystallization from isooctane. Proceed-
ing with (S)-6, we tried to prepare N-Boc-isofagomine (7)
according to a described procedure (Scheme 1).4 How-
ever, in our hands, two steps8 involving the incorporation
of the exocyclic methylene carbon were troublesome, ren-
dering this synthesis unsuitable for upscaling. To circum-
vent these problems we devised an alternative synthesis,9
which is depicted in the retrosynthetic scheme below
(Scheme 2). The piperidine derivative 9 is prepared via
alkylation of 2-benzyloxymethylallylamine (8) with (S)-
vinyloxirane [(S)-4], followed by RCM. In our approach,
all carbon atoms of isofagomine (1) are already present
prior to RCM. After suitable hydroxyl protection in 9 and
stereoselective hydroboration, the three hydroxyl groups
in 10 are differentiated.
OH
OH
OH
O
OH
OH
O
NH
HO
HO
HO
OH
OH
OH
OH
1
2
3
Figure 1
Syntheses of isofagomine (1) are numerous,1,2 starting
either from the chiral pool or from a prochiral precursor.
In most cases, however, enantiomer or diastereomer sep-
aration is cumbersome. In order to turn isofagomine (1)
into a synthetically useful and flexible chiral building
block, an efficient synthesis should provide reasonable
amounts of a suitably protected derivative, containing
three differentiated hydroxyl functions. Most of the hith-
erto reported syntheses of isofagomine (1) lack this possi-
bility for ready differentiation of the hydroxyl groups.
SYNLETT 2008, No. 15, pp 2321–2325
Advanced online publication: 22.08.2008
DOI: 10.1055/s-2008-1078280; Art ID: G08808ST
1
5
.0
9
.2
0
0
8
© Georg Thieme Verlag Stuttgart · New York

2322
P. E. R. Espeel et al.
LETTER
OH OBn
OBn
PGO
HO
O
+
OBn NH2
N
N
Boc
Boc
10
9
(S)-4
8
Scheme 2 Retrosynthesis
The large-scale preparation of 2-benzyloxymethylallyl-
amine (8) involved a six-step sequence with an overall
yield of 41% (Scheme 3). 3-Chloro-2-chloromethyl-1-
propene (11) was converted into 2-methylidene-1,3-pro-
panediol (12) via substitution with acetate and subsequent
methanolysis.10 Transformation to the cyclic sulfite 13
followed by treatment with sodium azide yielded 2-azi-
domethylprop-2-en-1-ol (14),11 which was converted into
2-benzyloxymethylallylamine (8)12 after O-benzylation
and Staudinger reduction.
Figure 2 Chiral HPLC analysis: Chiralpak AD-H; n-hexane–EtOH,
95:5 (isocratic); T = 35 °C; UV detection at l = 214 nm
9-BBN in different solvents and at elevated temperature
appeared to be very sluggish, due to steric hindrance.
Switching to the less sterically demanding TBDMS group
rendered 18 reactive towards hydroboration with thexyl-
borane (20; Scheme 5).12,16 Standard deprotection of hy-
droxyl and amine protecting groups in 19 afforded the
hydrochloride of isofagomine (1·HCl; 53% overall yield
from (S)-4].
d
a, b
e, f
c
68%
90%
86%
Cl
Cl
OH OH
O
O
OH N3 78%
OBn NH2
S
11
12
13
14
8
O
Scheme 3 Reagents and conditions: (a) AcOH, Et3N, reflux, over-
night; (b) K2CO3, MeOH, r.t., overnight; (c) SOCl2, CCl4, 0 °C, 50
An analogous hydroboration–deprotection sequence,
min; (d) NaN3, DMF, 80 °C, 15 min; (e) (i) NaH, DMF, 0 °C, 30 min; starting from 17, yielded (2S,3R,4R)-2,4-bis(hydroxy-
methyl)-3-hydroxypyrrolidine hydrochloride (24),15,17
novel C6 pyrrolidine azasugar [8% overall yield from (S)-
4]. The concomitant removal of the TBDMS group during
hydrogenolysis of 22 is noteworthy (Scheme 6).18
a
(ii) BnBr, DMF, r.t., overnight; (f) (i) Ph3P, THF, r.t., overnight; (ii)
2 M NaOH, 80 °C, 1 h.
Opening of the epoxide in (S)-4 by nucleophilic attack of
8 and subsequent amine protection led to a regioisomeric
mixture 15/16 (ratio 7:3), which could be separated
(Scheme 4).12,13 Products arising from an SN2¢ mechanism
were not detected. Both alkylation products 15 and 16
underwent RCM upon treatment with the Grubbs second
generation catalyst,14 yielding six- and five-membered
ring products 912 and 17,15 respectively, with more than
99% ee (Figure 2). This indicates that even in the case of
16, no racemization occurred, and a clean inversion of
configuration was assumed.
1
The H NMR and 13C NMR data as well as the optical
rotation19 {[a]D +20 (c = 0.93, EtOH)} confirmed the
structure of the synthesized isofagomine hydrochloride
(1·HCl). The structure of 24 was harder to prove. Because
of the conformational flexibility of five-membered rings,
1
the H–1H vicinal coupling constants of the ring protons
were useless for stereochemical assignment. Therefore,
the rigid bicyclic oxazolidinone 2520 was synthesized21
and submitted to a NOESY experiment: three pairs of nOe
contacts supported the anticipated stereochemistry
(Scheme 7).
In an attempt to obtain stereoselective hydroboration, we
initially chose to protect the hydroxyl function in 9 with
the bulky TBDPS group. However, hydroboration using
Azasugar 24 was tested for inhibition of several commer-
cial glycosidases22 (Table 1) and proved to be a slightly
OH OH
HO
OBn
OH OBn
OBn
b
HO
RO
TBDMSO
c, d, e
quant.
92%
b
N
BnO
NBoc OH
OBn NH2
Boc
93%
N
15 (64%)
9
N
N
H⋅HCl
a
Boc
Boc
8
+
BnO
+
1⋅HCl
19
9 R = H
b
a
OH
O
18 R = TBDMS 97%
90%
N
Boc
HO
BnO
NBoc
17
BH2
(S)-4
16 (16%)
20
Scheme 4 Reagents and conditions: (a) (i) 8 (3 equiv), H2O, 100 °C,
6 h; (ii) Boc2O, 1 M NaOH, dioxane–H2O, r.t., overnight; (b) Grubbs
second generation catalyst (3.25 mol%), CH2Cl2, r.t., 48 h.
Scheme 5 Reagents and conditions: (a) TBDMSCl, imidazole,
DMAP, CH2Cl2, r.t., overnight; (b) 20 (5 equiv), THF, 0 °C → r.t., 48
h, 30% aq H2O2, 2 M NaOH, 0 °C, 2 h; (c) TBAF, THF, r.t., 2 h; (d)
H2, Pd/C (10%), EtOH, r.t., overnight; (e) 1 M HCl, r.t., overnight.
Synlett 2008, No. 15, 2321–2325 © Thieme Stuttgart · New York

LETTER
Synthesis of Isofagomine and a New C6 Pyrrolidine Azasugar
2323
Acknowledgment
BnO
BnO
HO
FWO-Vlaanderen, the Federal Government [IAP-Phase VI (2007-
2011), Project P6/19] and Ghent University are greatly acknowl-
edged for financial support. P. Espeel is indebted to FWO-Vlaande-
ren for a research assistant position.
b
c
86%
77%
N
N
Boc
Boc
TBDMSO
22
RO
17 R = H
a
21 R = TBDMS 86%
References and Notes
(1) Stütz, A. E. Iminosugars as Glycosidase Inhibitors; Wiley-
VCH: Weinheim, 1999.
(2) Compain, P.; Martin, O. R. Iminosugars; Wiley: Chichester,
HO
HO
HO
7
HO
3
2007.
4
5
d
(3) Dong, W.; Jespersen, T.; Bols, M.; Skrydstrup, T.; Sierks,
M. R. Biochemistry 1996, 35, 2788.
(4) Ouchi, H.; Mihara, Y.; Takahata, H. J. Org. Chem. 2005, 70,
5207.
2
1
quant.
6
N
N
Boc
H.HCl
HO
HO
24
23
(5) [a]D +20 (c = 1, pentane).
(6) Schaus, S.; Brandes, B.; Larrow, J.; Tokunaga, M.; Hansen,
K.; Gould, A.; Furrow, M.; Jacobsen, E. J. Am. Chem. Soc.
2002, 124, 1307.
Scheme 6 Reagents and conditions: (a) TBDMSCl, imidazole,
DMAP, CH2Cl2, r.t., overnight; (b) 20 (5 equiv), THF, 0 °C → r.t.,
48 h, 30% aq H2O2, 2 M NaOH, 0 °C, 2 h; (c) H2, Pd/C (10%), EtOH,
r.t., 48 h; (d) 1 M HCl, r.t., overnight.
(7) Chiral HPLC analysis: Chiralcel OD-H; n-hexane–EtOH,
99:1 (isocratic); T = 35 °C; UV (l = 214 nm).
(8) Incomplete stereoselective epoxidation4 and incomplete
regioselective epoxide opening with vinyl cuprate4 both
yielded an inseparable mixture of product and starting
material.
BnO
nOe
Ha
HO
Hb
OH
H
H
N
a, b
3
1
6
2
O
35%
N
4
(9) For a recent related approach towards racemic isofagomine
(rac-1), see: Imahori, T.; Ojima, H.; Tateyama, H.; Mihara,
Y.; Takahata, H. Tetrahedron Lett. 2008, 49, 265.
(10) Jautelat, M.; Artl, D.; Wieland, H. Eur Patent, EP 0110245,
1984.
OBn
Boc
22
TBDMSO
5
O
25
Scheme 7 Reagents and conditions: (a) TBAF, THF, r.t., 2 h; (b)
diethylaminosulfur trifluoride (DAST), CH2Cl2, 0 °C→ r.t., 4 h.
(11) Friedrich, M.; Savchenko, A.; Wächtler, A.; de Meijere, A.
Eur. J. Org. Chem. 2003, 11, 2138.
(12) Selected Experimental Details:
Table 1 Glycosidase Inhibitor Activities (Ki [mM] at 37 °C)
O-Benzylation and Staudinger Reduction of 14: To an
ice-cooled solution of 14 (12.17 g, 107.6 mmol, colorless
oil) in anhyd DMF (180 mL) was added NaH (6.45 g, 60%
dispersion on mineral oil, 161.3 mmol). After stirring for 30
min at 0 °C, benzyl bromide (25.6 mL, 214 mmol) was
added at 0 °C and the reaction was stirred overnight (0 °C →
r.t.). NaOH (60 mL, 2 M aq solution) was added and the
mixture was stirred at 80 °C during 1 h. The reaction mixture
was diluted with H2O (1 L) and extracted with Et2O (3 × 1
L). The organic phase was washed with brine (1 L) and dried
(Na2SO4). The drying agent was filtered and the resulting
clear solution was evaporated under reduced pressure. The
oil was filtered over silica gel (n-hexane–EtOAc, 9:1) and
after evaporation, the crude residue was dissolved in anhyd
THF (570 mL). Ph3P (56.4 g, 215.2 mmol) was added and
the mixture was stirred overnight at r.t. NaOH (190 mL, 2 M
aq solution) was added and the mixture was refluxed for 1 h.
The mixture was diluted with H2O (1 L) and extracted with
CH2Cl2 (3 × 1 L). The organic phase was washed once with
brine (0.6 L) and dried (MgSO4). The drying agent was
filtered and the resulting clear solution was evaporated under
reduced pressure. After flash column chromatography on
silica gel and purification (EtOAc–CH2Cl2, 8:2 + 5% Et3N),
2-benzyloxymethylallylamine (8; 15 g, 84.63 mmol, 78%)
was isolated as a colorless oil.
Enzyme
(source)
a-Glucosidase b-Glucosidase a-Mannosidase
(yeast)
(almond)
(jack bean)
isofagomine 1
azasugar 24
rac-24
86
0.11
770
65
195
no inhibition
no inhibition
149
>1000
stronger inhibitor for a-glucosidase than isofagomine (1),
but a weaker inhibitor for b-glucosidase.3 No inhibition
was observed for a-mannosidase.23 The higher Ki values
of the racemate rac-24 indicate that, of the two enan-
tiomers, the synthesized (2S,3R,4R)-24 is the more potent
inhibitor.
In summary, enantiopure isofagomine (1) was synthe-
sized with an overall yield of 53%, starting from (S)-
vinyloxirane [(S)-4]. Key steps included nucleophilic
opening of (S)-vinyloxirane [(S)-4], ring-closing metathe-
sis and stereoselective hydroboration. The regioisomeric
alkylation product 16 of the epoxide opening was used in
an analogous reaction sequence yielding (2S,3R,4R)-2,4-
bis(hydroxymethyl)-3-hydroxypyrrolidine (24; 8% over-
all yield from (S)-4]. This novel C6 pyrrolidine azasugar is
a moderate inhibitor of a-glucosidase from yeast.
8: 1H NMR (300 MHz, CDCl3): d = 7.18–7.30 (m, 5 H), 5.03
(d, 2 H), 4.43 (s, 2 H), 3.97 (s, 2 H), 3.28 (s, 2 H), 1.21 (s, 2
H). 13C NMR (75 MHz, CDCl3): d = 147.6 (C), 138.3 (C),
128.4 (CH), 127.7 (CH), 127.7 (CH), 111.3 (CH2), 72.1
(CH2), 72.1 (CH2), 44.8 (CH2). HRMS (ES): m/z [M + H]+
calcd for C11H15NO: 178.12263; found: 178.12260.
Synlett 2008, No. 15, 2321–2325 © Thieme Stuttgart · New York

2324
P. E. R. Espeel et al.
LETTER
silica gel (n-hexane–EtOAc, 4:1), affording 19 (488 mg,
Alkylation of 2-Benzyloxymethylallylamine (8) with (S)-
Vinyloxirane [(S)-4]: 2-Benzyloxymethylallylamine (8;
5.24 g, 29.56 mmol) was mixed with H2O (132 mL, 7.33
mmol) in a pressure tube at 0 °C. (S)-Vinyloxirane [(S)-4;
0.8 mL, 9.93 mmol] was slowly added at 0 °C and the
reaction vessel was sealed, heated and stirred for 6 h at 100
°C. The reaction mixture was transferred to a 100-mL round-
bottomed flask and dissolved in H2O (5.8 mL), dioxane (35
mL) and NaOH (23 mL, 1 M in H2O). Boc2O (9.7 g, 44.44
mmol) was added at r.t. and the reaction mixture was stirred
overnight at r.t. The mixture was evaporated under reduced
pressure, diluted with Et2O (500 mL), subsequently washed
with H2O (200 mL), citric acid (200 mL, 20%) and brine
(200 mL). The pooled aqueous phases were extracted with
Et2O (3 × 250 mL). The organic phases were dried (MgSO4).
The drying agent was filtered and the resulting clear solution
was evaporated under reduced pressure. The residue (13 g)
was purified by flash column chromatography on silica gel
(gradient elution: n-hexane–EtOAc, 85:15 → 6:4) to furnish
15 (2.21 g, 6.35 mmol, 64%) and 16 (558 mg, 1.61 mmol,
16%) as colorless oils.
1.08 mmol, 93%) as a clear colorless oil.
19: [a]D –5 (c = 0.97, CHCl3). 1H NMR (300 MHz, CDCl3):
d = 7.20–7.30 (m, 5 H), 4.45 (s, 2 H), 4.06 (m, 2 H), 3.53 (br
s, 2 H), 3.34 (m, 2 H), 2.54 (m, 3 H), 1.76 (m, 1 H), 1.39 (s,
9 H), 0.84 (s, 9 H), 0.06 (s, 6 H). 13C NMR (75 MHz,
CDCl3): d = 154.6 (C), 138.2 (C), 128.4 (CH), 127.6 (CH),
127.5 (CH), 80.0 (C), 75.7 (CH), 73.3 (CH2), 73.1 (CH),
69.4 (CH2), 48.8 (CH2), 45.1 (CH2), 41.6 (CH), 28.4 (Me),
25.8 (Me), 18.1 (C), –4.4 (Me). HRMS (ES): m/z [M + Na]+
calcd for C24H41NO5Si: 452.28265; found: 452.28284
(13) Reported yields are isolated yields. The ratio of 15/16 was
determined by RP-HPLC: 30% → 65% MeCN in H2O in 60
min; Phenomenex Luna C18 (2), 5 m, 250 × 4.6 mm.
(14) RCM can also be accomplished by the Grubbs–Hoveyda
catalyst, however, not by the Grubbs first generation
catalyst.
(15) Solutions of Boc-protected 17, 21, 22 and 23 consisted of
rotamers complicating interpretation of NMR data.
(16) (a) Negishi, E.; Brown, H. C. Synthesis 1974, 77.
(b) Freshly prepared (according to the Aldrich Technical
Bulletin AL-109) 0.5 M solution of thexylborane in THF.
(17) 24: [a]D –4.0 (c = 0.72, H2O). 1H NMR (700 MHz, D2O):
d = 4.09 (app. t, J2,3 = J3,4 = 8.0 Hz, 1 H, H-3), 3.98 (dd,
15: [a]D 0 (c = 1, CHCl3). 1H NMR (300 MHz, CDCl3):
d = 7.18–7.30 (m, 5 H), 5.75 (ddd, J = 5.6, 10.5, 17.1 Hz, 1
H), 5.25 (d, J = 17.1 Hz, 1 H), 5.08 (m, 2 H), 4.94 (s, 1 H),
4.42 (s, 2 H), 4.26 (m, 1 H), 3.87 (m, 4 H), 3.58 (br s, 1 H),
3.21 (m, 2 H), 1.36 (s, 9 H). 13C NMR (75 MHz, CDCl3):
d = 157.5 (C), 141.9 (C), 138.4 (CH), 138.0 (C), 128.4 (CH),
127.7 (CH), 115.7 (CH2), 113.4 (CH2), 80.6 (C), 72.7 (CH),
72.2 (CH2), 71.3 (CH2), 53.7 (CH2), 51.5 (CH2), 28.3 (Me).
HRMS (ES): m/z [M + Na]+ calcd for C20H29NO4:
J2,6a = 3.6 Hz, J6a,6b = 12.6 Hz, 1 H, H-6a), 3.86 (dd, J2,6b
6.4 Hz, J6a,6b = 12.6 Hz, 1 H, H-6b), 3.82 (dd, J4,7a = 4.6 Hz,
J7a,7b = 11.6 Hz, 1 H, H-7a), 3.73 (dd, J4,7b = 6.3 Hz, J7a,7b
=
=
11.6 Hz, 1 H, H-7b), 3.66 (dd, J4,5a = 8.7 Hz, J5a,5b = 12.2
Hz, 1 H, H-5a), 3.58 (ddd, J2,6a = 3.6 Hz, J2,6b = 6.4 Hz, J2,3
= 8.0 Hz, 1 H, H-2), 3.25 (dd, J4,5b = 9.3 Hz, J5a,5b = 12.2 Hz,
1 H, H-5b), 2.48 (m, 1 H, H-4). 13C NMR (75 MHz, D2O): d
= 70.9 (C-3), 65.3 (C-2), 59.5 (C-7), 57.8 (C-6), 46.2 (C-4),
45.3 (C-5). IR (HATR): 3304, 2930, 2749, 1595, 1400,
1346, 1055, 1021, 956, 820, 640 cm–1. HRMS (ES): m/z [M
+ H]+ calcd for C6H13NO3: 148.09681; found: 148.09671.
(18) (a) When 19 was submitted to the same hydrogenolysis
conditions, the TBDMS protecting group remained intact.
(b) Ikawa, T.; Sajiki, H.; Hirota, K. Tetrahedron 2004, 60,
6189. (c) Hattori, K.; Sajiki, H.; Hirota, K. Tetrahedron
2001, 57, 2109.
370.19889; found: 370.19967.
16: [a]D +9.1 (c = 1, CHCl3). 1H NMR (300 MHz, CDCl3):
d = 7.18–7.30 (m, 5 H), 5.78 (m, 1 H), 5.04–5.15 (m, 4 H),
4.43 (s, 2 H), 4.26 (m, 1 H), 3.93 (m, 2 H), 3.80 (m, 2 H),
3.70 (m, 2 H), 1.37 (s, 9 H). 13C NMR (75 MHz, CDCl3):
d = 156.3 (C), 143.0 (C), 137.9 (C), 133.9 (CH), 128.4 (CH),
127.8 (CH), 117.5 (CH2), 113.9 (CH2), 80.4 (C), 72.3 (CH2),
71.7 (CH2), 63.5 (CH2), 62.0 (CH), 48.7 (CH2), 28.4 (Me).
HRMS (ES): m/z [M + Na]+ calcd for C20H29NO4:
370.19889; found: 370.19897.
Ring-Closing Metathesis of 15: To a solution of 15 (1.89 g,
5.44 mmol) in anhyd and degassed CH2Cl2 (170 mL) was
added the Grubbs second generation catalyst (150 mg, 0.177
mmol) at r.t. under an argon atmosphere and the solution was
stirred for 48 h. The solvent was evaporated and purified by
flash column chromatography on silica gel (n-hexane–
EtOAc, 2:1; silica gel saturated with Et3N) to furnish 9 (1.60
g, 5 mmol, 92%) as a brown solid.
(19) Jespersen, T.; Bols, M.; Sierks, M. R.; Skrydstrup, T.
Tetrahedron 1994, 50, 13449.
(20) 25: 1H NMR (700 MHz, CDCl3): d = 7.18–7.30 (m, 5 H, Ph),
4.46 (s, 2 H, OBn), 4.41 (dd, J = 7.4, 9.2 Hz, 1 H, H-1b), 4.27
(dd, J = 2.5, 9.2 Hz, 1 H, H-1a), 3.75 (m, 2 H, H-2, H-3), 3.62
(dd, J = 5.2, 9.0 Hz, 1 H, H-6b), 3.40 (m, 2 H, H-5a, H-6a),
3.22 (dd, J = 8.2, 11.7 Hz, 1 H, H-5b), 2.70 (br s, 1 H, OH),
2.45 (m, 1 H, H-4). 13C NMR (75 MHz, CDCl3): d = 161.4
(C), 137.4 (C), 128.7 (CH), 128.1 (CH), 127.8 (CH), 78.9
(CH), 73.7 (CH2), 71.5 (CH2), 66.3 (CH2), 64.1 (CH), 47.2
(CH), 46.7 (CH2).
9: [a]D +52.2 (c = 1.01, CHCl3). 1H NMR (300 MHz,
CDCl3): d = 7.19–7.30 (m, 5 H), 5.84 (m, 1 H), 4.43 (s, 2 H),
4.16 (br s, 1 H), 3.93 (d, J = 18.0 Hz, 1 H), 3.89 (s, 2 H), 3.73
(d, J = 18.1 Hz, 1 H), 3.46 (br s, 2 H), 1.40 (s, 9 H). 13C NMR
(75 MHz, CDCl3): d = 155.2 (C), 137.9 (C), 136.6 (C), 128.5
(CH), 127.8 (CH), 127.8 (CH), 125.4 (CH), 80.2 (C), 72.4
(CH2), 71.5 (CH2), 63.7 (CH), 47.7 (CH2), 44.0 (CH2), 28.4
(Me). HRMS (ES): m/z [M + Na]+ calcd for C18H25NO4:
342.16759; found: 342.16788.
Hydroboration of 18: Compound 18 (500 mg, 1.15 mmol)
was dissolved in anhyd THF (2 mL) and a freshly prepared
solution of thexylborane (20;16 5.77 mmol, 11.5 mL, 0.5 M
in THF) was added at 0 °C and the reaction was stirred for
48 h (0 °C → r.t.). The reaction was quenched by adding
30% H2O2–2 M NaOH (1:1; 8 mL) at 0 °C and the mixture
was then stirred for 2 h at 0 °C. The reaction was diluted with
brine (100 mL) and extracted with CH2Cl2 (3 × 100 mL).
After drying (Na2SO4), filtration and evaporation of the
solvent, the residue was purified by flash chromatography on
(21) Zhao, H.; Thurkauf, A. Synlett 1999, 1280.
(22) The inhibition constants (Ki) were determined using four
inhibitor concentrations within a limited range around the Ki
value.
Inhibition of b-Glucosidase (Almond): Ki was determined
at 37 °C using a NaH2PO4–Na2HPO4 buffer (pH 6.5; 100
mM)and 2-chloro-4-nitrophenyl-b-D-glucoside assubstrate.
The release of 2-chloro-4-nitrophenol was monitored
continuously by measuring absorbance at l = 405 nm. The
Ki values were determined by Dixon plots.
Inhibition of a-Glucosidase (Yeast): Ki was determined at
37 °C using a NaH2PO4–Na2HPO4 buffer (pH 5.6; 100 mM)
and 4-nitrophenyl-a-D-glucoside as substrate. The release of
4-nitrophenol was monitored by measuring absorbance at
l = 405 nm after addition of 10% Na2CO3 to samples of the
reaction mixture at regular time intervals. The Ki values were
Synlett 2008, No. 15, 2321–2325 © Thieme Stuttgart · New York

LETTER
determined by Dixon plots.
Synthesis of Isofagomine and a New C6 Pyrrolidine Azasugar
2325
mannoside as substrate, and were monitored analogously to
the a-glucosidase assay above.
(23) No significant inhibition of the glycolysis of 4-nitrophenyl-
a-D-mannoside was observed at a concentration of 1.9 mM
in the presence of 24 or rac-24 (1 mM).
Inhibition of a-Mannosidase (Jack Bean): Reactions were
performed at 37 °C using a HOAc–NaOAc buffer (pH 4.5;
100 mM) containing ZnCl2 (2 mM) and 4-nitrophenyl-a-D-
Synlett 2008, No. 15, 2321–2325 © Thieme Stuttgart · New York

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