Synthesis and glycosidase inhibition studies of 5-methyl-substituted tetrahydroxyindolizidines and -pyrrolizidines related to natural hyacinthacines B

Article abstract of DOI:10.1002/ejoc.201300103

The synthesis of three tetrahydroxyindolizidines and one tetrahydroxypyrrolizidine related to natural hyacinthacines B and their biological evaluation as glycosidase inhibitors is reported. The target molecules were obtained through highly stereoselective cycloadditions between sugriched allylic and homoallylic alcohols. This allowed the installation of a methyl group at C5 - a common feature of many natural hyacinthacines - with high control over the stereoselectivity. The new compounds inhibit amyloglucosidase from Aspergillus niger and β-glucosidase from almonds. Compound 1 is a competitive inhibitor of amyloglucosidase and shows a fair selectivity towards this enzyme. The presence of C5-Me substitution in indolizidines 2 and 3 slightly diminishes the inhibitory activity towards amyloglucosidase whereas it improves the inhibitory properties towards β-glucosidase. Cycloaddition between a carbohydrate-derived nitrone and suitable chemoenzymatically prepared enantioenriched allylic and homoallylic alcohols allowed the straightforward synthesis of three tetrahydroxyindolizidines and one tetrahydroxypyrrolizidine related to natural hyacinthacines B. They inhibit amyloglucosidase from Aspergillus niger and β-glucosidase from almonds. Copyright

Full text of DOI:10.1002/ejoc.201300103

FULL PAPER
DOI: 10.1002/ejoc.201300103
Synthesis and Glycosidase Inhibition Studies of 5-Methyl-Substituted
Tetrahydroxyindolizidines and -pyrrolizidines Related to Natural
Hyacinthacines B
Daniele Martella,[a,b] Francesca Cardona,[a] Camilla Parmeggiani,[a,c] Francisco Franco,[b]
Juan A. Tamayo,*[b] Inmaculada Robina,[d] Elena Moreno-Clavijo,[d]
Antonio J. Moreno-Vargas,[d] and Andrea Goti*[a]
Dedicated to Marek Chmielewski on the occasion of his 70th birthday
Keywords: Total synthesis / Azasugars / Alkaloids / Enzyme catalysis / Inhibitors / Cycloaddition
The synthesis of three tetrahydroxyindolizidines and one tet-
rahydroxypyrrolizidine related to natural hyacinthacines B
compounds inhibit amyloglucosidase from Aspergillus niger
and β-glucosidase from almonds. Compound 1 is a competi-
and their biological evaluation as glycosidase inhibitors is re- tive inhibitor of amyloglucosidase and shows a fair selectivity
ported. The target molecules were obtained through highly
stereoselective cycloadditions between sugriched allylic and
homoallylic alcohols. This allowed the installation of a methyl
group at C5 – a common feature of many natural hyacinthac-
ines – with high control over the stereoselectivity. The new
towards this enzyme. The presence of C5-Me substitution in
indolizidines 2 and 3 slightly diminishes the inhibitory ac-
tivity towards amyloglucosidase whereas it improves the in-
hibitory properties towards β-glucosidase.
Introduction
Hyacinthacines are polyhydroxylated pyrrolizidine alka-
loids bearing hydroxymethyl groups at C3 and in several
cases also methyl or hydroxymethyl groups at C5. Although
they were discovered only recently, around twenty of these
compounds were identified during the period 1999–2007,
by isolation from the Hyacinthaceae family of plants.[1]
They have been classified as A, B, or C on the basis of their
degrees of oxygenation (three, four, or five hydroxy groups,
respectively, Figure 1).
[a] Dipartimento di Chimica “Ugo Schiff”, Università di Firenze,
Polo Scientifico e Tecnologico,
Via della Lastruccia 3–13, 50019 Sesto Fiorentino (FI), Italy
Fax: +39-055-4573531
E-mail: andrea.goti@unifi.it
[b] Departamento de Química Farmacéutica y Orgánica, Facultad
de Farmacia, Universidad de Granada,
Figure 1. Examples of hyacinthacine alkaloids and the structurally
related steviamine.
Campus de la Cartuja s/n., 18071 Granada, Spain
E-mail: jtamayo@ugr.es
[c] CNR-INO and European Laboratory for Non-Linear
Spectroscopy (LENS), University of Florence,
Via N. Carrara 1, Sesto Fiorentino (FI), Italy
[d] Departamento de Química Orgánica, Facultad de Química,
Universidad de Sevilla,
Only in 2010 was the first example of an indolizidine
analogue bearing a methyl group at C5 – namely (–)-stevi-
amine – identified, in Stevia rebaudiana Betoni (Asteraceae)
and in Veltheimia capensis Hyacinthaceae (sand lily).[2]
c/ Prof. García González 1, 41012, Sevilla, Spain
Supporting information for this article is available on the
WWW under http://dx.doi.org/10.1002/ejoc.201300103.
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These natural iminosugars and their unnatural analogues sessing different configurations at the stereogenic centers.[6b]
behave as glycosidase inhibitors and so are promising can- This allowed control of the relative configurations at C2
didates as multipurpose drugs in the treatment of different and C3a of the isoxazolidine intermediates 5 and therefore
pathologies.[3] For this reason, synthetic efforts to develop at C7 (or C6 in the case of pyrrolizidines) and at the bridge-
general strategies to afford these compounds have been head carbon atoms in the target molecules. Finally, the rela-
enormous during recent years.[4] New synthetic methods to tive configuration at C5 depended on the enantiomer of the
access such compounds are important not only for confirm- alcohol 7, obtained through chemoenzymatic resolution
ation of the structures of the natural samples,[5] but also and used in the cycloaddition. Complete control of all
to provide new analogues for structure–activity relationship stereocenters could thus be achieved with high selectivity.
studies.
Our groups have a longstanding interest in the total syn-
Results and Discussion
thesis of pyrrolizidine and indolizidine alkaloids and ana-
logues.[6,7] In particular, Goti and co-workers have widely
exploited the use of polyhydroxylated nitrones as dipoles in
1,3-dipolar cycloaddition chemistry[6a,6b,4e] and as electro-
philes towards organometallics,[6c,6d] whereas Tamayo and
co-workers have recently expanded the scope of cycload-
ditions for the synthesis of unnatural pentahydroxypyrroliz-
idines and -indolizidines with hydroxymethyl groups at C5
by use of chemoenzymatically prepared but-3-ene-1,2-diol
derivatives as dipolarophiles.[8]
Here we report a general strategy for the synthesis of
novel tetrahydroxyindolizidines and -pyrrolizidines, there-
fore related to hyacinthacines B,[9] each bearing a methyl
group at C5 (see 2–4, Scheme 1). This is also a common
structural motif of these natural compounds (e.g., hya-
cinthacines A3, B3, B4, B7, C5, steviamine, Figure 1).[10] To
provide information about the role of the methyl group for
the inhibitory activity, model compound 1 was also synthe-
sized.
The first step for the synthesis of the model compound
1 involved cycloaddition between 6[11] and but-3-en-1-ol (8,
Scheme 2). Previous work in our group had shown that this
dipolarophile required prolonged reaction times with dif-
ferent cyclic nitrones,[12] but that these reaction times could
be greatly reduced (and stereoselectivity improved) under
microwave (MW) irradiation conditions.[13] We therefore
first attempted the reaction under microwave irradiation
conditions, with the aim of keeping the reaction times short.
However, the reaction between 6 and 8 under MW heating
conditions failed both in dichloromethane and in toluene
as solvents, affording only partial conversion of the starting
material. We then treated 6 with the O-acetyl derivative 9,
obtained from 8 by treatment with acetic anhydride in pyr-
idine at room temperature (see the Supporting Information)
and used without further purification. After MW heating
in toluene at 120 °C for two hours, the single adduct 10
(Scheme 2) was recovered in 81% yield after purification by
flash column chromatography (FCC). Removal of the
acetyl group was achieved by treatment with the strongly
basic resin Ambersep 900 OH at room temp. in MeOH.[6b]
Compound 11 was obtained by this two-step sequence in
78% overall yield. Alternatively, conventional heating of 6
with 8 in toluene at 60 °C for 4 d directly afforded 11 as a
single isomer in 89% yield. The structures of 10 and 11,
confirmed after subsequent transformation (vide infra),
confirmed our expectation that both cycloadditions pro-
ceeded through an exo approach of 8 or 9 anti to the vicinal
OBn of the nitrone. This stereochemical outcome is in ac-
cord with other reactions between nitrone 6 and different
dipolarophiles.[6a,14]
Scheme 1. General strategy for the synthesis of tetrahy-
droxyindolizidine and -pyrrolizidine alkaloids.
The general strategy is outlined in Scheme 1. Target com-
pounds 1–4 were obtained through highly regio- and stereo-
selective cycloadditions between a d-arabino-configured
nitrone 6 and a suitable allylic or homoallylic alcohol 7 as
the key step. We chose nitrone 6 for two main reasons:
firstly, many hyacinthacines (e.g., hyacinthacines A2, A3, B4,
C5, Figure 1) have a common structural motif represented
by a pyrrolidine ring with three contiguous stereocenters
with the R configuration, which corresponds to a d-arabino
Scheme 2. Synthesis of isoxazolidines 10 and 11. Reaction condi-
tions: a) toluene, MW, 120 °C, 2 h, 81%; b) toluene, 60 °C, 4 d,
89%; c) Ambersep 900 OH, MeOH, 96%.
Two different strategies to afford the indolizidine skel-
substitution pattern, and secondly, the use of nitrone 6 al- eton were investigated, as outlined in Scheme 3. Mesylation
lowed very high regio- and stereoselectivity in the cycload- of the primary alcohol in 11 (MsCl, triethylamine) gave the
dition reaction, which is not the case with nitrones pos- intermediate salt 12, formed by intramolecular mesylate
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Glycosidase Inhibition Studies
substitution with the nitrogen atom.[12a,15] Catalytic hydro-
To access the indolizidines with methyl substitution at
genolysis under acidic conditions resulted in N–O bond C5, the enantiomeric pent-4-en-2-ols were required as di-
cleavage and concomitant removal of the benzyl groups, polarophiles. Chemoenzymatic kinetic resolution of racemic
leading directly to the target compound 1 in 37% yield over pent-4-en-2-ol (15, Scheme 4) was achieved by stirring in
two steps.
diethyl ether at room temperature for 14 h in the presence
of Candida antarctica lipase Chirazyme L-2, c.-f., C2 and
vinyl acetate as acyl donor, as reported for different racemic
alcohols.[16] Under these conditions, acetate (R)-16 was re-
covered in 47% yield and 86% ee,[17] together with unre-
acted (S)-15 in 50% yield. The ee of unreacted 15 was deter-
mined after separation from (R)-16 by FCC and acetylation
in Ac2O and Py, which afforded (S)-16 quantitatively and
with 82% ee (Scheme 4).[17]
Scheme 3. Synthesis of tetrahydroxyindolizidine 1. Reaction condi-
tions: a) MsCl, NEt3, dry CH2Cl2; b) H2, Pd/C, MeOH, HCl, 37%
over two steps; c) I2, ImH, PPh3, dry THF, Na2S2O3, 35%; d) H2,
Pd/C, MeOH, HCl, 84%; e) I2, ImH, PPh3, dry THF, Na2S2O3,
then Ac2O/Py; f) H2, Pd/C, MeOH, HCl, 81% yield from 11.
Scheme 4. Chemoenzymatic preparation of (R)-16 and (S)-16 from
racemic 15. Reaction conditions: a) Chirazyme L-2, c.-f., C2, vinyl
acetate, diethyl ether, room temp., 14 h; b) Ac2O, Py, room temp.
Alternatively, treatment of the primary alcohol in 11 with
iodine and imidazole (ImH) in the presence of PPh3, fol-
lowed by Na2S2O3 treatment, resulted in a quite unexpected
outcome: indolizidine 13 was obtained in 35% yield. At this
stage, a plausible explanation for the formation of 13 in-
volved intramolecular nucleophilic displacement of the in-
termediate iodide by the nitrogen atom, followed by N–O
bond cleavage occurring during the reductive quenching
with Na2S2O3. To the best of our knowledge, however, there
is no report of such cleavage in similar isoxazolidines or
their salt derivatives. Subsequent catalytic hydrogenolysis
gave 1 in 84% yield. Because purification of 13 from the
triphenylphosphane oxide formed in the reaction of 11 was
troublesome, we decided to perform direct acetylation of
13 to afford 14, which was directly subjected to catalytic
hydrogenolysis under acidic conditions, leading to 1 in 81%
yield over three steps from 11.
Cycloaddition between 6 and enantioenriched (R)-16 in
toluene under microwave irradiation conditions at 120 °C
for three hours afforded the isoxazolidine 17 (Scheme 5) re-
gio- and stereoselectively in 74% yield, as a consequence of
the preferred exo approach of the dipolarophile anti with
respect to the vicinal OBn of the nitrone. The structure of
1
17 was assigned on the basis of its H NMR, 13C NMR,
COSY, HSQC, and 2D NOESY spectra. In particular, the
NOESY spectrum showed strong correlation peaks (Fig-
ure 3) between H(2) and H(6) and between H(3a) and H(5).
Treatment of 17 with the strongly basic resin Ambersep
900 OH in MeOH at room temp. led to 18 in 82% yield.
Treatment of the secondary alcohol with I2, imidazole, and
The structure of 1, which also confirmed that of its pre-
cursor 11, was assigned on the basis of 1H NMR, 13C
NMR, COSY, HSQC, and 1D NOESY spectra of its tet-
raacetyl derivative, obtained by treatment of 1 with a mix-
ture of Ac2O and pyridine (see the Supporting Infor-
mation). In particular, 1D NOESY spectra showed strong
NOE correlation peaks between H(7) and H(8a) and be-
tween H(7) and one of the two hydrogen atoms at C5 (Fig-
ure 2).
Scheme 5. Synthesis of tetrahydroxyindolizidines 2 and 3. Reaction
conditions: a) (R)-16, toluene, MW, 120 °C, 3 h, 74% for 17, 51%
for 20; b) Ambersep 900 OH, MeOH, 82% for 18, 75% for 21; c) I2,
ImH, PPh3, dry THF, Na2S2O3, 64% for 19, 73% for 22; d) H2,
Pd/C, MeOH, HCl, 95% for 2, 70% for 3.
Figure 2. Diagnostic NOE correlation peaks in in tetraacetates of
indolizidines 1, 2, and in 3.
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PPh3 in dry THF directly gave the indolizidine 19 in 64% intermolecular iodide attack, to form isoxazolidine salts
yield. As described above, the N–O bond cleavage probably analogous to 12. Such strained intermediates, in the pres-
occurred during the reductive quenching with Na2S2O3. Fi- ence of excess PPh3, are prone to reductive N–O bond
nal catalytic hydrogenolysis of 19 furnished the indolizidine cleavage by Na2S2O3 during the quenching procedure, ulti-
2 in 95% yield (Scheme 5). The 2D NOESY spectrum of mately affording the observed indolizidines 19, 22, and 13,
the acetate of 2, obtained by treatment of 2 with a mixture respectively. Support for this hypothesis was gained by
of Ac2O and pyridine (see the Supporting Information), treatment of the indolizidine salt 12 under the same reac-
showed strong NOE correlation peaks (Figure 2) of the tion conditions (PPh3, I2, ImH, then Na2S2O3): the indoliz-
methyl group at C5 with H(7) and H(8a), suggesting the S idinol 13 was clearly detected in the crude reaction mixture,
configuration at C5. The formation of indolizidine 19 from whereas no reaction was observed on quenching only with
isoxazolidine 18 would therefore have occurred with a single Na2S2O3.
inversion of configuration at C2Ј, contrary to our above
hypothesis of intermediate iodide formation.
The synthesis of pyrrolizidines with methyl substitution
at C5 required enantioenriched but-3-en-2-ol derivatives as
dipolarophiles. Unfortunately, the chemoenzymatic kinetic
resolution of racemic but-3-en-2-ol (23, Scheme 6), per-
formed with Candida antarctica lipase Chirazyme L-2, c.-f.,
C2 in the presence of vinyl acetate as acyl donor, was less
enantioselective. The reaction required only 5 h of stirring
at room temp. in diethyl ether to achieve 50% conversion
and furnished acetate (R)-24 in 39% yield but only
52% ee,[19] together with unreacted (S)-23 in 43% yield
(Scheme 6). The ee of unreacted 23 was determined after
Figure 3. Diagnostic NOE correlation peaks in isoxazolidines 17,
20, and 25.
Cycloaddition between 6 and enantioenriched (S)-16 in separation from (R)-24 by FCC; acetylation in Ac2O and
toluene under MW conditions at 120 °C for three hours Py afforded (S)-24 in 100% yield but with a low 42% ee
gave isoxazolidine 20 in 51% yield, and this was deacety- (Scheme 6).[19] Use of different enzymes for the selective
lated by treatment with Ambersep 900 OH in MeOH to af- acetylation of racemic 23 was also tried,[20] but no better
ford 21 in 75% yield (Scheme 5). The structure of 20, origi- results were achieved. Moreover, attempted chemoenzym-
nating from an exo approach of the dipolarophile anti with atic deacetylation of the acetate of 23 with Chirazyme L-2,
respect to the vicinal OBn of the nitrone, was assigned on c.-f., C2 was unsuccessful.
the basis of a strong NOE correlation peak between H(2)
and H(6) in the 2D NOESY spectrum (Figure 3).
The similar results of the above cycloadditions with en-
antioenriched (R)-16 and (S)-16 demonstrated that an ad-
ditional stereogenic center at C2 of the dipolarophile has
virtually no influence on the stereoselectivity of the reac-
tions, which is governed by substrate control by the nitrone.
Treatment of 21 with I2, ImH, and PPh3, together with
reductive quenching conditions, afforded the indolizidine 22
in 73% yield from 21. While this manuscript was in prepa-
Scheme 6. Chemoenzymatic preparation of (R)-24 and (S)-24 from
racemic 23. Reaction conditions: a) Chirazyme L-2, c.-f., C2, vinyl
acetate, diethyl ether; b) Ac2O, Py, room temp.
ration, compound 22 was also synthesized through an imine
1
cycloaddition strategy.[18] The H and 13C NMR spectra of
22 were identical to those reported, thus demonstrating the
However, cycloaddition between 6 and an excess
R configuration at C5 unequivocally. Final catalytic hydro- (5 equiv.) of enantioenriched (R)-24 in toluene under MW
genolysis of 22 furnished the indolizidine 3 in 70% yield conditions at 120 °C for three hours afforded a reasonable
(Scheme 5).
61% yield of the major exo-anti diastereoisomer 25
As a further confirmation of the structures of the final (Scheme 7), which was isolated from the reaction mixture
tetrahydroxyindolizidines, comparison of the 1H NMR by FCC. The structure of 25 was assigned on the basis of
spectra of 2 and 3 showed a signal at around 3.3 ppm for a strong NOE correlation peak between H(2) and H(6) in
the equatorial H(5) in 2 and a more shielded signal at the 2D NOESY spectrum (Figure 3). Treatment with the
around 2.6 ppm for the axial H(5) in 3. The formation of strongly basic resin Ambersep 900 OH in MeOH at room
indolizidines 19 and 22 from isoxazolidines 18 and 21 thus temp. led to 26 in quantitative yield. Treatment of 26 with
occurred with a single inversion of configuration at C2Ј in I2, ImH, and PPh3 in THF or toluene, though, was unsuc-
each case. This result can be reasonably explained by as- cessful, even with prolonged reaction times or heating at
suming that treatment of cycloadducts 18, 21, and 11 with 60–80 °C. This observation confirmed the difficulties in-
an excess of I2 and PPh3 in the presence of imidazole does herent in the use of external iodide to effect displacement
not afford an intermediate iodide, but that intramolecular reactions with secondary alcohols of this kind. On the other
nucleophilic substitution by the nitrogen atom of the hy- hand, internal displacement by nitrogen is in this case pre-
droxy group activated by PPh3/I2 occurs more rapidly than cluded by the high strain of the putative product.
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Glycosidase Inhibition Studies
Table 1. Inhibitory activities of compounds 1, 2, 3, and 4 with
glycosidases. Percentage inhibition at 1 mm, IC50 (in parenthe-
sis, μm) and Ki (bold, μm) if measured. Optimal pH, 37 °C.[a,b,c]
α-l-Fuc-
α-Gal- Amylo Glc- α-Glc-
β-Glc-
β-Man-
ase
ase
ase
ase
ase
ase
1
32
n.i.
95 (20.6)
18 (C)
83
96 (35)
34.8
n.i.
21
15
2
3
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
88 (122) n.i.
87 (144) 18
4
n.i.
17
95 (39)
32.1
18
27
n.i.
[a] For conditions of measurements see ref.[21] and the Supporting
Information. [b] (C): competitive inhibition, n.i.: no inhibition at
1 mm concentration of the inhibitor. [c] α-l-Fuc-ase: α-l-fucosidase
from bovine kidney. α-Gal-ase: α-galactosidase from coffee beans.
AmyloGlc-ase: amyloglucosidase from Aspergillus niger. α-Glc-ase:
α-glucosidase from yeast. β-Glc-ase: β-glucosidase from almonds.
β-Man-ase: β-mannosidase from snail.
Scheme 7. Synthesis of tetrahydroxypyrrolizidine 4. Reaction con-
ditions: a) toluene, MW, 120 °C, 3 h, 61%; b) Ambersep 900 OH,
MeOH, 100%; c) MsCl, dry CH2Cl2, Py, 95%; d) Zn powder,
AcOH/H2O (9:1), 47%; e) H2, Pd/C, MeOH, HCl, 80%.
Compound 1 showed the best inhibitory activity towards
amyloglucosidase from Aspergillus niger with a competitive
type of inhibition (see the Supporting Information for Li-
neweaver–Burk plot). This compound also showed weak in-
hibitory properties with α-l-fucosidase, β-glucosidase, and
β-mannosidase. The presence of a methyl group at C5
somewhat decreased the inhibitory properties with amylog-
lucosidase from Aspergillus niger, as was also observed for
compounds 2, 3, and 4. The methyl group in compound 2
thus diminished the inhibitory potential towards amyloglu-
cosidase from Aspergillus niger and increased the activity
against β-glucosidase from almonds to IC50 = 122 μm. The
methyl groups in compounds 3 and 4 reduced the activity
to Ki 34.8 μm and Ki 32.1 μm, respectively. Indolizidine 3
also showed moderate to weak inhibitory activity with β-
glucosidase from almonds (IC50 = 144 μm), whereas pyrroli-
zidine 4 showed weak inhibitory properties. Compound 3
also exhibited weak inhibitory activity against β-mannos-
idase from snail. Compound 4 is also a weak inhibitor of
α-galactosidase and α- and β-glucosidases.
We then decided to obtain the pyrrolizidine skeleton in
a two-step sequence based on mesylation of the secondary
alcohol 26 followed by N–O bond cleavage. Accordingly, 26
was treated with mesyl chloride and pyridine in dry CH2Cl2
to afford compound 27 (Scheme 7) in 95% yield. Subse-
quent treatment with Zn in an AcOH/H2O mixture (9:1)
gave the pyrrolizidine 28 in 47% yield. Compound 28 was
assigned the S configuration at C5, as the result of a single
inversion of configuration occurring during the intramolec-
ular nucleophilic substitution by the nitrogen atom after N–
O bond cleavage. This assignment was supported by the
strong NOE correlation peak between H(5) and H(3) in the
2D NOESY spectrum of 28 (Figure 4). Final catalytic hy-
drogenolysis of 28 furnished the pyrrolizidine 4 in 80%
yield (Scheme 7).
In summary, the presence of a methyl group at C5 (com-
pounds 2, 3, and 4) slightly reduces the inhibitory activity
towards amyloglucosidase from Aspergillus niger. It does,
however, improve the inhibitory properties towards β-
glucosidase from almonds for indolizidines 2 and 3, but not
for pyrrolizidine 4.
Figure 4. Diagnostic NOE correlation peaks in pyrrolizidine 28.
Biological Evaluation of Compounds 1–4 with Glycosidases
Conclusions
Synthesized compounds 1–4 were assayed for their in-
hibitory activity towards 11 commercially available glycos-
In conclusion, the straightforward synthesis of three new
idases.[21] At 1 mm concentration and under enzyme-opti- compounds related to natural hyacinthacines B was
mal pH they did not inhibit β-galactosidase from Aspergil- achieved by means of highly regio- and stereoselective 1,3-
lus orizae or from Escherichia coli, α-glucosidase from rice, dipolar cycloadditions between a carbohydrate-derived
α-mannosidase from Jack beans, or β-N-acetylglucosamini- nitrone and suitable chemoenzymatically prepared enantio-
dase from Jack beans. Table 1 summarizes the inhibitory enriched allylic and homoallylic alcohols. This strategy al-
activities measured with α-l-fucosidase from bovine kidney, lowed stereoselective introduction of methyl substitution at
α-galactosidase from coffee beans, amyloglucosidase from C5 in the target molecules. Biological evaluation with a set
Aspergillus niger, α-glucosidase from yeast, β-glucosidase of commercially available glycosidases revealed that these
from almonds, and β-mannosidase from snail.
new compounds mainly inhibit amyloglucosidase form As-
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sep 900 (OH– form, 1.5 g) was added to a solution of 10 (435 mg,
0.82 mmol) in MeOH (75 mL), and the mixture was stirred over-
night. TLC monitoring (AcOEt/hexane 3:1) revealed a new slower-
running compound (Rf = 0.38). The mixture was filtered, washed
with MeOH, and concentrated to give 11 (386 mg, 96%) as a yellow
pergillus niger and β-glucosidase from almonds. The best
inhibitor in this series is compound 1, which preferentially
inhibits amyloglucosidase from Aspergillus niger. The
methyl substitution at C5 in indolizidines 2 and 3 slightly
diminishes inhibition of amyloglucosidase, but improves the
activity against β-glucosidase. Pyrrolizidine 4 has inhibitory
properties towards amyloglucosidase similar to those of
compound 3, although its activity against β-glucosidase is
lower.
1
oil. [α]2D5 = –36.3 (c = 1, CHCl3). H NMR (400 MHz, CDCl3): δ
= 7.36–7.24 (m, 15 H, Ar), 4.62–4.53 (m, 6 H, Bn), 4.32–4.25 (m,
1 H, 2-H), 4.04 (dd, J = 6.8, 4.9 Hz, 1 H, 5-H), 3.92 (t, J = 4.9 Hz,
1 H, 4-H), 3.77–3.71 (m, 3 H, 3a-H, CH2OH), 3.67 (dd, J = 9.8,
4.9 Hz, 1 H, CH2OBn), 3.62 (dd, J = 10.0, 5.6 Hz, 1 H, CH2OBn),
3.33 (dt, J = 5.9, 5.4 Hz, 1 H, 6-H), 2.21 (ddd, J = 12.2, 6.3, 3.9 Hz,
1 H, 3-Ha), 2.13–2.06 (m, 1 H, 3-Hb), 1.92–1.77 (m, 2 H,
CH2CH2OH) ppm. 13C NMR (100 MHz, CDCl3): δ = 140.9, 140.7,
140.4 (s, Ar), 131.1–130.2 (d, 15 C, Ar), 90.4 (d, C-4), 87.8 (d, C-
5), 77.6 (d, C-2), 76.1, 75.1, 74.6 (t, Bn), 72.4 (t, CH2OBn), 72.1
(d, C-6), 70.4 (d, C-3a), 63.0 (t, CH2OH), 42.8 (t, C-3), 38.3 (t,
Experimental Section
General Methods: Commercial reagents were used as received. All
reactions were magnetically stirred and monitored by TLC on
0.25 mm silica gel plates (Merck F254); column chromatography
was carried out on silica gel 60 (32–63 μm), yields refer to spectro-
scopically and analytically pure compounds unless otherwise
stated. Small-scale microwave-assisted synthesis was carried out
with an Initiator 2.0 single-mode microwave instrument producing
controlled irradiation at 2.450 GHz (Biotage AB, Upsala). These
parameters were established by the basic principles of microwave-
assisted organic synthesis. The temperature was measured with an
IR sensor on the outside of the reaction vessel. NMR spectra were
recorded with Varian INOVA-400, Bruker AMX-300, or Bruker
ARX-400 instruments. Infrared spectra were recorded with a Per-
kin–Elmer FTIR Spectrum One spectrophotometer. Mass spectra
were recorded with Micromass Model Platform II and Autospec-Q
instruments; relative percentages are shown in brackets. Elemental
analyses were performed with a Perkin–Elmer 2400 analyzer. Op-
tical rotation measurements were performed with a JASCO DIP-
370 polarimeter. GLC was performed with a gas chromatograph
equipped with split/splitless injector and a flame-ionization detec-
tor. The He flow rate was 1:1 mLmin–1, the injection port and the
zone-detector temperatures were 275 °C; β-DEXTM 325 (Supelco
TM) capillary column (30ϫ0.25 mm i.d.ϫ0.25 mm film thickness)
at 80 °C.
CH CH OH) ppm. IR (CHCl ): ν = 3538, 3032, 2868, 2248, 1496,
˜
2
2
3
1454, 1364 cm–1 . HRMS (ES): calcd. for C30 H3 5NO5 Na
[M + Na]+ 512.2413; found 512.2426.
(2R,3aR,4R,5R,6R)-4,5-Dibenzyloxy-6-(benzyloxymethyl)-2-(2-
hydroxyethyl)-hexahydropyrrolo[1,2-b]isoxazole (11): A solution of
6 (150 mg, 0.36 mmol) and but-3-en-1-ol (153 μL, 1.79 mmol) in
toluene (1.5 mL) was heated at 60 °C. After 48 h the solvent was
removed and the residue was flash chromatographed (AcOEt/petro-
leum ether 2:1, Rf = 0.32) to afford 11 (156 mg, 0.32 mmol, 89%)
as a yellow oil.
(1R,2R,3R,7S,8aR)-1,2-Dibenzyloxy-7-hydroxy-3-(benzyloxymeth-
yl)indolizidine (13): Iodine (179 mg, 0.69 mmol), triphenylphos-
phane (161 mg, 0.61 mmol), and imidazole (96 mg, 1.14 mmol)
were added under Ar to a stirred solution of 11 (230 mg,
0.47 mmol) in dry THF (30 mL). The mixture was stirred at room
temperature for 2 h. TLC monitoring (AcOEt/hexane 3:1) showed
the appearance of a new product (Rf = 0.21). A saturated aqueous
solution of NaHCO3 and Na2S2O3 (20 mL) was added, and the
mixture was stirred at room temperature for 15 min. It was then
extracted with CH2Cl2 (3ϫ 100 mL). The combined organic phase
was dried with Na2SO4, filtered, and concentrated under reduced
pressure. The residue was flash chromatographed (AcOEt/hexane
4:1, Rf = 0.18) and then retained on a column of Dowex 50WX8
(200–400 mesh). The column was washed with MeOH (100 mL),
water (100 mL), and then with NEt3 (10%) in MeOH to afford
pure 13 (78 mg, 35%) as a yellow oil. [α]2D5 = –20.1 (c = 1, CHCl3).
1H NMR (400 MHz, CDCl3): δ = 7.34–7.28 (m, 15 H, Ar), 4.58–
4.46 (m, 6 H, Bn), 3.89 (t, J = 2.7 Hz, 1 H, 2-H), 3.70 (dd, J = 4.6,
2.6 Hz, 1 H, 1-H), 3.63–3.57 (m, 2 H, 7-H, CH2OBn), 3.50 (dd, J
= 9.5, 6.0 Hz, 1 H, CH2OBn), 3.36–3.34 (m, 1 H, 3-H), 3.10–3.08
(m, 1 H, 5-Ha), 2.98–2.96 (m, 1 H, 8a-H), 2.67 (td, J = 12.0, 2.0 Hz,
1 H, 5-Hb), 2.02–2 (m, 1 H, 8-Ha), 1.78–1.76 (m, 1 H, 6-Ha), 1.53
(qd, J = 11.9, 4.2 Hz, 1 H, 6-Hb), 1.38 (q, J = 11.5 Hz, 1 H, 8-
Hb) ppm. 13C NMR (100 MHz, CDCl3): δ = 132.1, 132.0, 131.9 (s,
Ar), 128.8–127.6 (d, 15 C, Ar), 88.1 (d, C-1), 86.6 (d, C-2), 73.3,
71.7, 71.6 (t, Bn), 69.3 (t, CH2OBn), 69.2 (d, C-7), 64.4 (d, C-3),
62.7 (d, C-8a), 44.7 (t, C-5), 37.6 (t, C-8), 32.3 (t, C-6) ppm. IR
(CHCl ): ν = 3379, 3059, 2923, 2852, 1590, 1437 cm–1. HRMS
(2R,3aR,4R,5R,6R)-2-(2-Acetyloxyethyl)-4,5-dibenzyloxy-6-(benz-
yloxymethyl)-hexahydropyrrolo[1,2-b]isoxazole (10): A solution of
nitrone 6 (495 mg, 1.19 mmol) and 9 (1.35 mg, 11.8 mmol) in tolu-
ene (2 mL) was heated at 120 °C under microwave irradiation con-
ditions in a sealed tube. After 2 h, TLC (AcOEt/hexane 3:1) re-
vealed the presence of a new compound of higher Rf (= 0.74). The
solvent was removed, and the residue was flash chromatographed
(AcOEt/hexane 1:2, Rf = 0.22) to afford 10 (629 mg, 81%) as a
brown oil. [α]2D7 = –31.9 (c = 1, CHCl3). 1H NMR (400 MHz,
CDCl3): δ = 7.36–7.24 (m, 15 H, Ar), 4.63–4.54 (m, 6 H, Bn), 4.18–
4.15 (m, 2 H, 2-H, CH2OAc), 4.13–4.08 (m, 1 H, CH2OAc), 4.05
(t, J = 5.7 Hz, 1 H, 5-H), 3.91 (t, J = 4.8 Hz, 1 H, 4-H), 3.76 (br.
s, 1 H, 3a-H), 3.69 (dd, J = 9.7, 4.1 Hz, 1 H, CH2OBn), 3.63 (dd,
J = 9.7, 4.8 Hz, 1 H, CH2OBn), 3.30 (br. s, 1 H, 6-H), 2.21–2.17
(m, 1 H, 3-Ha), 2.06–2.04 (m, 1 H, 3-Hb), 2.02 (s, 3 H, CH3CO),
1.96–1. 93 (m, 1 H, CH2 CH2 OAc), 1. 89–1.85 (m, 1 H,
CH2CH2OAc) ppm. 13C NMR (100 MHz, CDCl3): δ = 173.6 (s,
CH3CO), 140.9, 140.6, 140.4 (s, Ar), 131.1–130 (d, 15 C, Ar), 90.2
(d, C-4), 85.6 (d, C-5), 76.1, 75.2, 74.6 (d, t, 4 C, C-2, Bn), 72.4 (t,
CH2OBn), 72.1 (d, C-6), 70.5 (d, C-3a), 64.3 (t, CH2OAc), 42.9 (t,
˜
3
[nanostructure-assisted laser desorption/ionization time of flight
(NALDI-TOF)]: calcd. for C30H36NO4 [M + H]+ 474.2644; found
474.2649.
C-3), 35.0 (t, CH CH OAc), 23.6 (q, CH CO) ppm. IR (CHCl ): ν
˜
2
2
3
3
(1R,2R,3R,7S,8aR)-1,2,7-Trihydroxy-3-(hydroxymethyl)indolizid-
ine (1): A solution of 13 (73 mg, 0.15 mmol) in MeOH (15 mL) was
acidified (conc. HCl, five drops) and hydrogenated (60 psi H2) in
= 2935, 2866, 1738, 1454, 1365, 1242 cm–1. HRMS (ES): calcd. for
C32H37NO6Na [M + Na]+ 554.2519; found 554.2516.
(2R,3aR,4R,5R,6R)-4,5-Dibenzyloxy-6-(benzyloxymethyl)-2-(2- the presence of Pd/C (10%, 30 mg) for 72 h. The catalyst was fil-
hydroxyethyl)- hexahydropyrrolo[1,2-b]isoxazole (11): Amber- tered off, washed with MeOH, and concentrated to give a residue
4052
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© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2013, 4047–4056

Glycosidase Inhibition Studies
that was retained on a column of Dowex 50WX8 (200–400 mesh).
The column was thoroughly washed with MeOH (75 mL), water
(75 mL) and then with NH4OH (1 m, 100 mL) to afford pure 1
(26 mg, 84%) as a viscous yellow oil. [α]2D4 = +2.2 (c = 0.91,
idine (5 mL) and Ac2O (4.7 mL) gave (S)-2-O-acetyl-pent-4-en-2-ol
[(S)-16, 2.67 g, 100%, tR = 10.90 min; 82% ee].
(2R,2ЈR,3aR,4R,5R,6R)-2-(2-Acetyloxypropyl)-4,5-dibenzyloxy-
6-(benzyloxymethyl)-hexahydropyrrolo[1,2-b]isoxazole (17): A solu-
tion of 6 (471 mg, 1.13 mmol) and (R)-16 (1.44 g, 11.3 mmol) in
toluene (2 mL) was heated at 120 °C under microwave irradiation
conditions in a sealed tube. After 3 h, TLC (AcOEt/hexane 3:1)
revealed the presence of a new compound (Rf = 0.79). The solvent
was removed, and the residue was flash chromatographed (AcOEt/
hexane 1:2, Rf = 0.30) to afford 17 (456 mg, 74%) as a yellow oil.
1
MeOH), H NMR (400 MHz, D2O): δ = 3.81 (t, J = 4.8 Hz, 1 H,
2-H), 3.69 (dd, J = 11.8, 5.0 Hz, 1 H, CH2OH), 3.65–3.59 (m, 2 H,
1-H, CH2OH), 3.57–3.53 (m, 1 H, 7-H), 2.94–2.86 (m, 2 H, 5-Ha,
3-H), 2.68 (ddd, J = 11.5, 6.3, 3 Hz, 1 H, 8a-H), 2.55 (dt, J = 12.7,
2.9 Hz, 1 H, 5-Hb), 1.97–1.92 (m, 1 H, 8-Ha), 1.75–1.71 (m, 1 H,
6-Ha), 1.34 (qd, J = 12.5, 4.6 Hz, 1 H, 6-Hb), 1.15 (q, J = 11.7 Hz,
1 H, 8-Hb) ppm. 13C NMR (50 MHz, D2O): δ = 80.1 (d, C-1), 78.7
(d, C-2), 67.4 (d, C-7), 65.4 (d, C-3), 62.5 (d, C-8a), 59.1 (t,
CH2OH), 42.7 (t, C-5), 34.6 (t, C-6), 30.1 (t, C-8) ppm. HRMS
(NALDI-TOF): calcd. for C9H18NO4 [M + H]+ 204.1236; found
204.1239.
[α]2D4 = –37.9 (c = 0.6, CHCl3). H NMR (500 MHz, CDCl3): δ =
1
7.32–7.25 (m, 15 H, Ar), 5.00–4.94 [m, 1 H, CH(CH3)OAc], 4.63–
4.52 (m, 6 H, Bn), 4.14–4.09 (m, 1 H, 2-H), 4.03 (dd, J = 6.7,
4.6 Hz, 1 H, 5-H), 3.90 (t, J = 4.6 Hz, 1 H, 4-H), 3.76–3.73 (m, 1
H, 3a-H), 3.70 (dd, J = 9.7, 4.1 Hz, 1 H, CH2OBn), 3.63 (dd, J =
9.7, 6.2 Hz, 1 H, CH2OBn), 3.29 (pseudo q, J = 5.6 Hz, 1 H, 6-H),
2.16 (ddd, J = 12.3, 6.2, 3.7 Hz, 1 H, 3-Ha), 2.05–2.01 (m, 1 H, 3-
Hb), 1.99 [s, 3 H, CH(CH3)OAc], 1.84–1.80 [m, 2 H, CH2CH(CH3)-
OAc], 1.23 (d, J = 6.3 Hz, 3 H, Me) ppm. 13C NMR (100 MHz,
CDCl3): δ = 170.4 (s, CH3CO) 138.6, 138.3, 138.1 (s, Ar), 128.5–
127.6 (d, 15 C, Ar), 88.0 (d, C-4), 83.4 (d, C-5), 73.6 (t, Bn), 73.1
(d, C-2) 72.6, 72.0 (t, Bn), 70.3 (t, CH2OBn), 69. 8 (d, C-6), 69.2
[d, CH(CH3)OAc], 68.1 (d, C-3a), 40.9 (t, C-3), 39.9 [t,
CH2CH(CH3)OAc], 21.3 (q, CH3CO), 20.6 [q, CH(CH3)OAc] ppm.
(1R,2R,3R,7S,8aR)-1,2,7-Trihydroxy-3-(hydroxymethyl)indolizid-
ine (1): NEt3 (73 μL, 0.52 mmol) was added under nitrogen to a
stirred solution of 11 (170 mg, 0.35 mmol) in dry CH2Cl2 (3 mL).
MsCl (30 μL, 0.38 mmol) was added dropwise at 0 °C. The solution
was stirred at room temp. for 45 min. TLC analysis (AcOEt/petro-
leum ether 3:1) showed disappearance of the starting material. The
solvent was removed, and the residue was dissolved in MeOH
(9 mL). The solution was acidified (conc. HCl, five drops) and hy-
drogenated in the presence of Pd/C (10%, 110 mg) for 72 h. The
catalyst was filtered off, with washing with MeOH, and the solu-
tion was concentrated to give a residue that was retained on a col-
umn of Dowex 50WX8 (200–400 mesh). The column was thor-
oughly washed with MeOH (20 mL), water (10 mL), and then with
NH4OH (6%, 40 mL) to afford pure 1 (25.9 mg, 0.13 mmol, 37%)
as a viscous yellow oil.
IR (CHCl ): ν = 3031, 2934, 2869, 1738, 1455, 1372, 1244 cm–1.
˜
3
HRMS (NALDI-TOF): calcd. for C33H40NO6 [M + H]+ 546.2856;
found 546.2861.
(2R,2ЈR,3aR,4R,5R,6R)-4,5-Dibenzyloxy-6-(benzyloxymethyl)-2-
(2-hydroxypropyl)-hexahydropyrrolo[1,2-b]isoxazole (18): Am-
bersep 900 (OH– form, 1.6 g) was added to a solution of 17
(456 mg, 0.84 mmol) in anhydrous MeOH (70 mL), and the mix-
ture was stirred overnight. TLC (AcOEt/hexane 3:1) revealed a new
slower-running compound (Rf = 0.67). The mixture was filtered
and washed with MeOH. The solvent was removed. The residue
was flash chromatographed (AcOEt/hexane 3:2, Rf = 0.45) to yield
18 (344 mg, 82%) as a yellow oil. [α]2D7 = –36.2 (c = 0.4, CHCl3).
1H NMR (400 MHz, CDCl3): δ = 7.26–7.18 (m, 15 H, Ar), 4.56–
4.46 (m, 6 H, Bn), 4.30–4.23 (m, 1 H, 2-H), 3.98 (dd, J = 6.8,
5.1 Hz, 1 H, 5-H), 3.94–3.89 [m, 1 H, CH(CH3)OH], 3.85 (t, J =
4.9 Hz, 1 H, 4-H), 3.69–3.65 (m, 1 H, 3a-H), 3.60 (dd, J = 9.9,
4.5 Hz, 1 H, CH2OBn), 3.55 (dd, J = 9.9, 5.3 Hz, 1 H, CH2OBn),
3.26 (dt, J = 11.4, 5.1 Hz, 1 H, 6-H), 2.11 (ddd, J = 12.4, 6.6,
3.9 Hz, 1 H, 3-Ha), 2.05–2.00 (m, 1 H, 3-Hb), 1.68–1.63 [m, 2 H,
CH2CH(CH3)OH], 1.10 [d, J = 6.5 Hz, 3 H, CH(CH3)OH] ppm.
13C NMR (100 MHz, CDCl3): δ = 141.0, 140.7, 140.5 (s, Ar),
131.1–130.2 (d, 15 C, Ar), 90.5 (d, C-4), 85.7 (d, C-5), 76.2 (d, C-
2), 76.0, 75.2, 74.6 (t, Bn), 72.4 (t, CH2OBn), 72.0 (d, C-6), 70.5
(d, C-3a), 67.6 [d, CH(CH3)OH], 44.3 [t, CH2CH(CH3)OH], 42.6
(1R,2R,3R,7S,8aR)-1,2,7-Trihydroxy-3-(hydroxymethyl)indolizid-
ine (1): Iodine (119 mg, 0.46 mmol), triphenylphosphane (104 mg,
0.40 mmol), and imidazole (63 mg, 0.92 mmol) were added under
N2 to a stirred solution of 11 (152 mg, 0.31 mmol) in dry THF
(20 mL). The mixture was stirred at room temp. for 2 h. TLC moni-
toring (AcOEt/hexane 3:1) showed the appearance of a new prod-
uct (Rf = 0.21). A saturated aqueous solution of NaHCO3 and
Na2S2O3 (20 mL) was added, and the mixture was stirred at room
temp. for 15 min. It was then extracted with CH2Cl2 (3ϫ 100 mL).
The combined organic phases were dried with Na2SO4, filtered,
and concentrated under reduced pressure. The residue was stirred
in the presence of acetic anhydride (3 mL) and dry pyridine
(12 mL) at room temp. for 18 h. The solvent was removed, and the
residue was purified by FCC (AcOEt/petroleum ether 1:2, Rf = 0.5)
to afford crude 14, which was dissolved in MeOH (9 mL). The
solution was acidified (conc. HCl, five drops) and hydrogenated in
the presence of Pd/C (10%, 60 mg) for 72 h. The catalyst was fil-
tered off and washed with MeOH, and the solution was stirred
for 14 h with Ambersep 900 (OH– form, 300 mg). The mixture was (t, C-3), 26.4 [q, CH(CH )OH] ppm. IR (CHCl ): ν = 3412, 3031,
filtered, washed with MeOH, and concentrated to give 1 (51 mg,
0.25 mmol, 81%) as a yellow oil.
˜
3
3
2931, 2866, 1455, 1103 cm–1. HRMS (NALDI-TOF): calcd. for
C31H38NO5 [M + H]+ 504.2750; found 504.2755.
Enzymatic Acetylation of Racemic Pent-4-en-2-ol: Chirazyme L-2,
c.-f., C2 (245 mg) was added to a gently stirred solution of (R,S)-
(1R,2R,3R,5S,7S,8aR)-1,2-Dibenzyloxy-3-(benzyloxymethyl)-7-
hydroxy-5-methylindolizidine (19): Iodine (221 mg, 0.87 mmol), tri-
pent-4-en-2-ol (3.5 g, 40.6 mmol) and vinyl acetate (3.76 mL, phenylphosphane (198 mg, 0.75 mmol), and imidazole (118 mg,
40.6 mmol) in Et2O (45 mL), and the mixture was stirred at room 1.74 mmol) were added under Ar to a stirred solution of 18
temp. The reaction was monitored by GLC analysis, and after 14 h
the enzyme was removed by filtration with thorough washing with
ether. After evaporation of the solvent, the residue was subjected
to chromatography (Et2O/pentane 1:2) to afford (R)-2-O-acetyl-
pent-4-en-2-ol [(R)-16, 2.45 g, 47 %, Rf = 0.65, tR = 12.92 min,
(292 mg, 0.58 mmol) in dry THF (25 mL). The mixture was stirred
at room temperature for 18 h. TLC monitoring (AcOEt/hexane 3:1)
showed the appearance of a new product (Rf = 0.46). A saturated
aqueous solution of NaHCO3 and Na2S2O3 (20 mL) was added,
and the mixture was stirred at room temperature for 15 min. It was
then extracted with CH2Cl2 (3ϫ 100 mL). The combined organic
fractions were dried with Na2SO4, filtered, and concentrated under
86% ee] and (S)-pent-4-en-2-ol [(S)-15, 1.7 g, 50%, Rf = 0.25, tR
=
6.58 min]. Conventional acetylation of (S)-15 in anhydrous pyr-
Eur. J. Org. Chem. 2013, 4047–4056
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
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4053

J. A. Tamayo, A. Goti et al.
FULL PAPER
reduced pressure. The residue was flash chromatographed (AcOEt/
ture was stirred overnight. TLC (AcOEt/hexane 3:1) revealed a new
hexane 2:1, Rf = 0.3) to afford 19 (181 mg, 64%) as a yellow oil slower-running compound (Rf = 0.69). The mixture was filtered
still containing some impurities. 1H NMR (400 MHz, CDCl3): δ =
and washed with MeOH. The solvent was removed, and the residue
7.36–7.26 (m, 15 H, Ar), 4.70–4.56 (m, 6 H, Bn), 3.94–3.89 (m, 1
was flash chromatographed (AcOEt/hexane 3:2, Rf = 0.46) to yield
H, 7-H), 3.85 (br., 1 H, 2-H), 3.78–3.75 (m, 1 H, 1-H), 3.61–3.51 21 (147 mg, 75%) as a yellow oil. [α]2D8 = –40.3 (c = 0.35, CHCl3).
(m, 2 H, CH2OBn), 3.43–3.41 (m, 2 H, 5-H, 8a-H), 3.29–3.27 (m, 1H NMR (400 MHz, CDCl3): δ = 7.36–7.25 (m, 15 H, Ar), 4.62–
1 H, 3-H), 1.68–1.52 (m, 4 H, 6-Ha, 6-Hb, 8-Ha, 8-Hb) 1.19 (d, J = 4.52 (m, 6 H, Bn), 4.28–4.23 (m, 1 H, 2-H), 4.06 (dd, J = 7.0,
6.2 Hz, 3 H, Me) ppm. 13C NMR (100 MHz, CDCl3): δ = 141.1, 5.0 Hz, 1 H, 5-H), 3.96–3.91 [m, 2 H, 4-H, CH(CH3)OH], 3.75–
141.0, 140.9 (s, Ar), 131.1–130.2 (d, 15 C, Ar), 89.5 (d, C-1), 89.5 3.72 (m, 1 H, 3a-H), 3.67 (dd, J = 9.5, 4.4 Hz, 1 H, CH2OBn), 3.63
(d, C-2), 75.9 (t, Bn), 75.1 (t, CH2OBn), 74.2, 73.9 (t, Bn), 67.8 (d, (dd, J = 9.5, 5.1 Hz, 1 H, CH2OBn), 3.34–3.31 (m, 1 H, 6-H), 2.20
C-3), 67.6 (d, C-7), 61.3 (d, C-8a), 51.9 (d, C-5), 37.2 (t, 2 C, C-6,
C-8), 23.0 (q, Me) ppm. HRMS (NALDI-TOF): calcd. for
C31H38NO4 [M + H]+ 488.2801; found 488.2804.
(ddd, J = 12.3, 6.2, 3.8 Hz, 1 H, 3-Ha), 2.05 (dt, J = 12.3, 8.8 Hz,
1 H, 3-Hb), 1.73–1.70 [m, 2 H, CH2CH(CH3)OH], 1.19 [d, J =
6.0 Hz, 3 H, CH(CH3)OH] ppm. 13C NMR (100 MHz, CDCl3): δ
= 140.9, 140.7, 140.4 (s, Ar), 131.1–130.1 (d, 15 C, Ar), 90.3 (d, C-
4), 85.6 (d, C-5), 78.2 (d, C-2), 76.1, 75.2, 74.6 (t, Bn), 72.3 (t,
CH2OBn), 72.2 (d, C-6), 70.1 (d, C-3a), 69.7 [d, CH(CH3)OH], 45.0
[t, CH2CH(CH3)OH], 43.5 (t, C-3), 23.0 [q, CH(CH3)OH] ppm. IR
(1R,2R,3R,5S,7S,8aR)-1,2,7-Trihydroxy-3-hydroxymethy-5-methyl-
indolizidine (2): A solution of 19 (98 mg, 0.20 mmol) in MeOH
(15 mL) was acidified (conc. HCl, five drops) and hydrogenated
(60 psi H2) in the presence of Pd/C (10%, 30 mg) for 72 h. The
catalyst was filtered off, washed with MeOH, and concentrated to
give a residue that was retained on a column of Dowex 50WX8
(200–400 mesh). The column was thoroughly washed with MeOH
(75 mL), water (75 mL), and then with NH4OH (1 m, 100 mL) to
afford pure 2 (42 mg, 95%) as a viscous yellow oil. [α]3D1 = –25.5 (c
(CHCl ): ν = 3430, 2931, 2866, 1455, 1104 cm–1. HRMS (NALDI-
˜
3
TOF): calcd. for C31H38NO5 [M + H]+ 504.2750; found 504.2749.
(1R,2R,3R,5R,7S,8aR)-1,2-Dibenzyloxy-3-(benzyloxymethyl)-7-
hydroxy-5-methylindolizidine (22): Iodine (111 mg, 0.44 mmol), tri-
phenylphosphane (99 mg, 0.38 mmol), and imidazole (59 mg,
0.87 mmol) were added under Ar to a stirred solution of 21
(147 mg, 0.29 mmol) in dry THF (15 mL). The mixture was stirred
at room temperature for 18 h. TLC monitoring (AcOEt/hexane 3:1)
showed the appearance of a new product (Rf = 0.47). A saturated
aqueous solution of NaHCO3 and Na2S2O3 (20 mL) was added,
and the mixture was stirred at room temperature for 15 min. It was
then extracted with CH2Cl2 (3ϫ 100 mL). The combined organic
fractions were dried with Na2SO4, filtered, and concentrated under
reduced pressure. The residue was purified by FCC (AcOEt/hexane
2:1, Rf = 0.21) to afford pure 22 (102 mg, 73%) as a yellow oil.
1
= 1.1, MeOH). H NMR (400 MHz, CD3OD): δ = 3.90–3.87 (m,
1 H, 7-H), 3.85–3.84 (m, 1 H, 1-H), 3.72 (dd, J = 5.5, 3.5 Hz, 1 H,
2-H), 3.65 (dd, J = 11.2, 4.3 Hz, 1 H, CH2OH), 3.60 (dd, J = 11.2,
4.0 Hz, 1 H, CH2OH), 3.31–3.29 (m, 1 H, 5-H), 3.18–3.15 (m, 1 H,
8a-H), 2.96 (pseudo q, J = 4.3 Hz, 1 H, 3-H), 1.70 (dtd, J = 12.8,
4.2, 1.5 Hz, 8-Ha), 1.64–1.60 (m, 1 H, 6-Ha), 1.56–1.52 (m, 2 H, 6-
Hb, 8-Hb), 1.16 (d, J = 6.8 Hz, 3 H, Me) ppm. 13C NMR
(100 MHz, CD3OD): δ = 84.0 (d, C-1), 83.4 (d, C-2), 70.9 (d, C-
3), 66.6 (d, C-7), 64.1 (t, CH2OH), 63.9 (d, C-8a), 50.9 (d, C-5),
36.4 (t, C-6), 36.1 (t, C-8), 21.7 (q, Me) ppm. HRMS (NALDI-
TOF): calcd. for C10H20NO4 [M + H]+ 218.1392; found 218.1399.
[α]2D7 = –28.2 (c = 0.35, CHCl3). H NMR (400 MHz, CDCl3): δ =
1
7.28–7.18 (m, 15 H, Ar), 4.62–4.38 (m, 6 H, Bn), 3.90–3.88 (m, 1
H, 2-H), 3.66–3.65 (m, 1 H, 1-H), 3.63–3.60 (m, 1 H, CH2OBn),
3.52–3.47 (m, 2 H, 3-H, 7-H), 3.41–3.37 (m, 1 H, CH2OBn), 2.68–
2.64 (m, 1 H, 8a-H), 2.59–2.55 (m, 1 H, 5-H), 2.19–2.16 (m, 1 H,
8-Ha), 1.81–1.78 (m, 1 H, 6-Ha), 1.20–1.13 (m, 2 H, 6-Hb, 8-Hb),
1.10 (d, J = 6.0 Hz, 3 H, Me) ppm. 13C NMR (100 MHz, CDCl3):
δ = 144.2, 144.0, 143.9 (s, Ar), 134.1–133.3 (d, 15 C, Ar), 95.1 (d,
C-1), 91.8 (d, C-2), 79.3, 77.7, 77.1 (t, Bn), 74. 6 (d, C-7), 71.4 (t,
CH2OBn), 68.2 (d, C-8a), 68.0 (d, C-3), 55.1 (d, C-5), 49.2 (t, C-
(2R,2ЈS,3aR,4R,5R,6R)-2-(2-Acetyloxypropyl)-4,5-dibenzyloxy-
6-(benzyloxymethyl)-hexahydropyrrolo[1,2-b]isoxazole (20): A solu-
tion of 6 (318 mg, 0.76 mmol) and (S)-16 (0.97 g, 7.6 mmol) in tolu-
ene (1.5 mL) was heated at 120 °C under microwave irradiation
conditions in a sealed tube. After 3 h, TLC (AcOEt/hexane 3:1)
revealed the presence of a new compound of higher Rf (0.67). The
solvent was removed, and the residue was flash chromatographed
(AcOEt/hexane 1:2, Rf = 0.32) to afford 20 (415 mg, 51%) as a
brown oil. [α]2D6 = –21.9 (c = 0.5, CHCl3). 1H NMR (400 MHz,
CDCl3): δ = 7.37–7.24 (m, 15 H, Ar), 5.02–4.98 [m, 1 H, CH(CH3)-
OAc], 4.63–4.52 (m, 6 H, Bn), 4.13–4.10 (m, 1 H, 2-H), 4.03 (dd,
J = 7.5, 4.7 Hz, 1 H, 5-H), 3.88 (t, J = 4.6 Hz, 1 H, 4-H), 3.73 (br.
s, 1 H, 3a-H), 3.68 (dd, J = 9.4, 4.7 Hz, 1 H, CH2OBn), 3.62 (dd,
J = 9.5, 4.8 Hz, 1 H, CH2OBn), 3.29–3.26 (m, 1 H, 6-H), 2.17 (ddd,
J = 12.2, 5.9, 3.4 Hz, 1 H, 3-Ha), 2.07–2.01 (m, 1 H, 3-Hb), 1.98
(s, 3 H, CH3CO), 1.96–1.93 [m, 1 H, CH2CH(CH3)OAc], 1.71 [ddd,
J = 14.0, 6.3, 4.8 Hz, 1 H, CH2CH(CH3)OAc], 1.24 [d, J = 5.7 Hz,
3 H, CH(CH3)OAc] ppm. 13C NMR (100 MHz, CDCl3): δ = 173.2
(s, CH3CO) 141.0, 140.7, 140.4 (s, Ar), 131.1–130.1 (d, 15 C, Ar),
90.3 (d, C-4), 85.3 (d, C-5), 76.08 (t, Bn), 75.4 (d, C-2) 75.3, 74.6
(t, Bn), 72.4 (t, CH2OBn), 71.8 (d, C-6), 71.3 [d, CH(CH3)OAc],
70.2 (d, C-3a), 43.4 (t, C-3), 41.7 [t, CH2CH(CH3)OAc], 24.0 (q,
6), 45.5 (t, C-8), 25.8 (q, Me) ppm. IR (CHCl ): ν = 3401, 2930,
˜
3
2865, 1367, 1455 cm–1. HRMS (NALDI-TOF): calcd. for
C31H38NO4 [M + H]+ 488.2801; found 488.2805.
(1R,2R,3R,5R,7S,8aR)-1,2,7-Trihydroxy-3-hydroxymethyl-5-
methylindolizidine (3): A solution of 22 (92 mg, 0.19 mmol) in
MeOH (15 mL) was acidified (conc. HCl, five drops) and hydroge-
nated (60 psi H2) in the presence of Pd/C (10%, 30 mg) for 72 h.
The catalyst was filtered off, washed with MeOH, and the solution
was concentrated to give a residue that was retained on a column
of Dowex 50WX8 (200–400 mesh). The column was thoroughly
washed with MeOH (75 mL), water (75 mL), and then with
NH4OH (1 m, 100 mL) to afford pure 3 (29 mg, 70%) as a viscous
1
yellow oil. [α]2D9 = –28.5 (c = 1.25, MeOH). H NMR (400 MHz,
CH CO), 23.0 [q, CH(CH )OAc] ppm. IR (CHCl ): ν = 3031, 2934,
˜
3
3
3
D2O): δ = 3.92–3.91 (m, 1 H, 2-H), 3.75 (dd, J = 11.7, 3.8 Hz, 1
H, CH2OH), 3.59–3.51 (m, 3 H, 1-H, 7-H, CH2OH), 2.97–2.95 (m,
1 H, 3-H), 2.67–2.64 (m, 1 H, 5-H), 2.59 (dt, J = 11.0, 2.1 Hz, 1
2860, 1738, 1455, 1373, 1247 cm–1. HRMS (NALDI-TOF): calcd.
for C33H40NO6 [M + H]+ 546.2856; found 546.2861.
(2R,2ЈS,3aR,4R,5R,6R)-4,5-Dibenzyloxy-6-(benzyloxymethyl)-2- H, 8a-H), 2.06–2.04 (m, 1 H, 8-Ha), 1.84–1.81 (m, 1 H, 6-Ha, 1.07–
(2-hydroxypropyl)-hexahydropyrrolo[1,2-b]isoxazole (21): Am-
bersep 900 (OH– form, 750 mg) was added to a solution of 20
(214 mg, 0.39 mmol) in anhydrous MeOH (70 mL), and the mix-
1.00 (m, 2 H, 6-Hb, 8-Hb), 0.99 (d, J = 5.9 Hz, 3 H, Me) ppm. 13C
NMR (100 MHz, D2O): δ = 84.6 (d, C-1), 82.4 (d, C-2), 70.3 (d,
C-7), 68.0 (d, C-3), 65. 7 (d, C-8a), 60.5 (t, CH2OH), 51.6 (d, C-
4054
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© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
Eur. J. Org. Chem. 2013, 4047–4056

Glycosidase Inhibition Studies
5), 44.0 (t, C-6), 39.5 (t, C-8), 21.1 (q, Me) ppm. HRMS (NALDI-
TOF): calcd. for C10H20NO4 [M + H]+ 218.1392; found 218.1385.
Pyridine (1.9 mL) and MsCl (51 μL, 0.64 mmol) were added to a
stirred solution of 26 (155 mg, 0.32 mmol) in anhydrous CH2Cl2
(7 mL). The reaction mixture was stirred at room temp. for 18 h.
TLC (AcOEt/hexane 3:1) showed the absence of 26 and the pres-
ence of a faster-moving compound (Rf = 0.84). MeOH (5 mL) was
added, and the reaction mixture was stirred for 20 min and then
concentrated to a residue that was subjected to FCC (AcOEt/hex-
ane 1:2) to give 27 (170 mg, 95%) as a white syrup. [α]3D1 = –14.4
Enzymatic Acetylation of Racemic But-3-en-2-ol: Chirazyme L-2,
c.-f., C2 (35 mg) was added to a gently stirred solution of (R,S)-
but-3-en-2-ol (1 g, 13.87 mmol) and vinyl acetate (0.64 mL,
6.9 mmol) in dry Et2O (8 mL), and the mixture was maintained
at room temp. with stirring. The reaction was monitored by GLC
analysis, and after 5 h the enzyme was removed by filtration, with
thorough washing with ether, and the solvent was evaporated. The
residue was subjected to FCC (Et2O/pentane 1:4) to afford (R)-2-
1
(c = 0.4, CHCl3). H NMR (400 MHz, CDCl3): δ = 7.37–7.25 (m,
15 H, Ar), 4.65 [pseudo quint, J = 6.6 Hz, 1 H, CH(CH3)OMs],
4.57–4.49 (m, 6 H, Bn), 4.20 (q, J = 7.3 Hz, 1 H, 2-H), 4.03 (dd, J
= 5.6, 4.3 Hz, 1 H, 5-H), 3.95 (t, J = 4.3 Hz, 4-H), 3.78–3.74 (m,
1 H, 3a-H), 3.64 (dd, J = 9.9, 5.4 Hz, 1 H, CH2OBn), 3.57 (dd, J
= 9.9, 5.7 Hz, 1 H, CH2OBn), 3.37 (pseudo q, J = 5.5 Hz, 1 H, 6-
H), 3.00 (s, 3 H, Ms), 2.25–2.08 (m, 2 H, 3-Ha, 3-Hb), 1.37 [d, J =
6.5 Hz, 3 H, CH(CH3)OMs] ppm. 13C NMR (100 MHz, CDCl3):
δ = 138.1, 137.8, 137.6 (s, Ar), 128.5–127.6 (d, 15 C, Ar), 87.4 (d,
C-4), 83.8 (d, C-5), 79.5 [d, CH(CH3)OMs], 78.4 (d, C-2), 73.4,
72.0, 72.3 (t, Bn), 70.1 (d, C-6), 69.9 (t, CH2OBn), 68.3 (d, C-3a),
38.3 (q, Ms) 36.4 (t, C-3), 18.7 [q, CH(CH3)OMs] ppm. IR
O-acetyl-but-3-en-2-ol [(R)-24, 611 mg, 39 %, Rf = 0.74, tR
=
6.22 min, 52% ee] and (S)- but-3-en-2-ol [(S)-23, 214 mg, 43%, Rf
= 0.22, tR = 3.84 min]. Conventional acetylation of (S)-23 (162 mg,
2.25 mmol) in anhydrous pyridine (1 mL) and Ac2O (318 μL) gave
(S)-2-O-acetyl-but-3-en-2-ol [(S)-24, 257 mg, 100%, tR = 6.11 min,
40% ee].
(1ЈR,2R,3aR,4R,5R,6R)-2-(1-Acetyloxyethyl)-4,5-dibenzyloxy-6-
(benzyloxymethyl)-hexahydropyrrolo[1,2-b]isoxazole (25): A solu-
tion of 6 (167 mg, 0.40 mmol) and (R)-24 (229 mg, 2 mmol) in tolu-
ene (0.8 mL) was heated at 120 °C under microwave irradiation
conditions in a sealed tube. After 3 h, TLC (AcOEt/hexane 3:1)
revealed the presence of a new compound of higher Rf (0.73). The
solvent was removed, and the residue was flash chromatographed
(AcOEt/hexane 1:4, Rf = 0.13) to yield 25 (130 mg, 61%) as a color-
less syrup. [α]2D7 = –18.6 (c = 0.4, CHCl3). 1H NMR (400 MHz,
CDCl3): δ = 7.37–7.28 (m, 15 H, Ar), 5.00–4.95 [m, 1 H, CH(CH3)-
OAc], 4.63–4.54 (m, 6 H, Bn), 4.23 (td, J = 7.6, 5.5 Hz, 1 H, 2-H),
4.07 (dd, J = 6.0, 3.1 Hz, 1 H, 5-H), 3.96 (pseudo t, J = 3.7 Hz, 1
H, 4-H), 3.76–3.74 (m, 2 H, 3a-H, CH2OBn), 3.65 (dd, J = 9.9,
6.5 Hz, 1 H, CH2OBn), 3.35 (td, J = 5.9, 5.3 Hz,1 H, 6-H), 2.16–
2.13 (m, 2 H, 3-Ha, 3-Hb), 2.05 (s, 3 H, CH3CO), 1.25 [d, J =
6.5 Hz, 3 H, CH(CH3)OAc] ppm. 13C NMR (100 MHz, CDCl3): δ
= 170.5 (s, CH3CO), 138.5, 138.1, 137.9 (s, Ar), 128.5–127.6 (d, 15
C, Ar), 87.2 (d, C-4), 83.3 (d, C-5), 77.9 (d, C-2) 73.5, 72.4, 71.9
(t, Bn), 70.5 [d, CH(CH3)OAc], 70.2 (t, CH2OBn), 69.9 (d, C-6),
68.3 (d, C-3a), 36.2 (t, C-3), 21.3 (q, CH3Ac), 16.5 [q, CH(CH3)-
(CHCl ): ν = 2925, 2856, 1454, 1357, 1101 cm–1. HRMS (NALDI-
˜
3
TOF): calcd. for C31H38NO7S [M + H]+ 568.2369; found 568.2360.
(1R,2R,3R,5S,6R,7aR)-1,2-Dibenzyloxy-3-(benzyloxymethyl)-6-
hydroxy-5-methylpyrrolizidine (28): A mixture of 27 (162 mg,
0.28 mmol) and Zn dust (148 mg) in AcOH/H2O 9:1 (3 mL) was
heated to 60 °C for 4 h. TLC monitoring (AcOEt/hexane 2:1)
showed the absence of 27 and the presence of a lower Rf (= 0.22)
compound. The reaction mixture was filtered through cotton with
washing with AcOEt. The solvent was washed with a saturated
solution of Na2CO3 and then evaporated. The residue was purified
by FCC (AcOEt/hexane 2:1) to afford 28 (63 mg, 47%). [α]2D7
=
+1.3 (c = 1.0, CHCl3). 1H NMR (400 MHz, CDCl3): δ = 7.36–7.25
(m, 15 H, Ar), 4.65–4.50 (m, 6 H, Bn), 4.13 (t, J = 4.3 Hz, 1 H, 2-
H), 4.06 (t, J = 4.3 Hz, 1 H, 1-H), 3.89 (q, J = 6.2 Hz, 1 H, 6-H),
3.61–3.56 (m, 1 H, 7a-H), 3.51 (d, J = 6.4 Hz, 2 H, CH2OBn), 3.38
(td, J = 6.4, 4.5 Hz, 1 H, 3-H), 3.01 (quint, J = 6.2 Hz, 1 H, 5-H),
2.36 (ddd, J = 12.8, 7.9, 6.3 Hz, 1 H, 7-Ha), 1.80 (dt, J = 12.9,
6.3 Hz, 1 H, 7-Hb), 1.12 (d, 3 H, Me) ppm. 13C NMR (100 MHz,
CDCl3): δ = 138.5 138.2, 138.0 (s, Ar), 128.4–127.5 (d, 15 C, Ar),
89.8 (d, C-1), 86.1 (d, C-2), 78.6 (d, C-6), 73.2, 72.1, 71.8 (t, 4 C,
Bn, CH2OBn), 69.3 (d, C-5), 68.9 (d, C-3), 66.2 (d, C-7a), 38.9 (t,
OAc] ppm. IR (CHCl ): ν = 2920, 2859, 1743, 1454, 1242 cm–1.
˜
3
HRMS (NALDI-TOF): calcd. for C32H38NO6 [M + H]+ 532.2699;
found 532.2703.
(1ЈR,2R,3aR,4R,5R,6R)-4,5-Dibenzyloxy-6-benzyloxymethyl-2-
(1-hydroxyethyl)-hexahydropyrrolo[1,2-b]isoxazole (26): Am-
bersep 900 (OH– form, 750 mg) was added to a solution of 25
(215 mg, 0.40 mmol) in anhydrous MeOH (70 mL), and the mix-
ture was stirred overnight. TLC (AcOEt/hexane 3:1) then revealed
a new slower-running compound (Rf = 0.48). The mixture was fil-
tered and washed with MeOH. The solvent was removed, and the
residue was purified by FCC (AcOEt/hexane 1:1, Rf = 0.21) to yield
26 (198 mg, 100%) as a colorless oil. [α]2D9 = –48.9 (c = 0.45,
CHCl3). 1H NMR (400 MHz, CDCl3): δ = 7.36–7.25 (m, 15 H,
Ar), 4.62–4.50 (m, 6 H, Bn), 4.07 (dd, J = 5.8, 4.3 Hz, 1 H, 5-H),
4.02 (td, J = 6.9, 5.8 Hz, 1 H, 2-H), 3.96 (pseudo t, J = 4.3 Hz, 1
H, 4-H), 3.77–3.74 (m, 1 H, 3a-H), 3.70–3.66 [m, 2 H, CH2OBn,
CH(CH3)OH], 3.62–3.59 (m, 1 H, 6-H), 3.38–3.35 (m, 1 H,
CH2OBn), 2.20–2.17 (m, 2 H, 3-Ha, 3-Hb), 1.18 [d, J = 6.4 Hz, 3
H, CH(CH3)OH] ppm. 13C NMR (100 MHz, CDCl3): δ = 140.9,
140.6, 140.4 (s, Ar), 131.1–130.2 (d, 15 C, Ar), 90.2 (d, C-4), 86.6
(d, C-5), 83.7 (d, C-2), 76.0, 75.0, 74.5 (t, Bn), 72.5 (d, C-6), 72.3
(t, CH2OBn), 71.9 [d, CH(CH3)OH], 71.2 (d, C-3a), 39.2 (t, C-3),
C-7), 19.5 (q, Me) ppm. IR (CHCl ): ν = 3396, 3063, 2865, 1454,
˜
3
1101 cm–1. HRMS (NALDI-TOF): calcd. for C30H36NO4
[M + H]+ 474.2644; found 474.2650.
(1R,2R,3R,5S,6R,7aR)-1,2,6-Trihydroxy-3-hydroxymethyl-5-
methylpyrrolizidine (4): A solution of 28 (59 mg, 0.12 mmol) in
MeOH (10 mL) was acidified (conc. HCl, five drops) and hydroge-
nated (60 psi H2) in the presence of Pd/C (10%, 25 mg) for 48 h.
The catalyst was filtered off, washed with MeOH, and the solution
was concentrated to give a residue that was retained on a column
of Dowex 50WX8 (200–400 mesh). The column was thoroughly
washed with MeOH (50 mL), water (50 mL), and then with
NH4OH (1 m, 75 mL) to afford pure 4 (20 mg, 80%) as a viscous
yellow oil. [α]2D4 = +7.0 (c = 0.70, H2O). 1H NMR (400 MHz, D2O):
δ = 3.87 (pseudo t, J = 8.1 Hz, 1 H, 1-H), 3.77 (q, J = 6.7 Hz; 1
H, 6-H), 3.69 (pseudo t, J = 8.1 Hz, 1 H, 2-H), 3.55 (dd, J = 11.7,
4.8 Hz, 1 H, CH2OH), 3.48 (dd, J = 11.7, 5.9 Hz, 1 H, CH2OH),
3.05 (td, J = 8.3, 5.9 Hz, 1 H, 7a-H), 2.83 (dt, J = 8.3, 5.4 Hz, 1
H, 3-H), 2.79–2.74 (m, 1 H, 5-H), 2.23 (ddd, J = 13.1, 7.9, 6.8 Hz,
1 H, 7-Ha), 1.68–1.62 (m, 1 H, 7-Hb), 0.94 (d, J = 6.4 Hz, 3 H,
Me) ppm. 13C NMR (50 MHz, D2O): δ = 80.5 (d, C-1), 77.8 (d, C-
22.3 [q, CH(CH )OH] ppm. IR (CHCl ): ν = 3437, 2929, 2867,
˜
3
3
1455, 1366, 1103 cm–1. HRMS (NALDI-TOF): calcd. for
C30H36NO5 [M + H]+ 490.2593; found 490.2597.
(1ЈR,2R,3aR,4R,5R,6R)-4,5-Dibenzyloxy-6-(benzyloxymethyl)-2-
(1-methylsulfonyloxyethyl)-hexahydropyrrolo[1,2-b]isoxazole (27): 2), 77.0 (d, C-6), 68.8 (d, C-3), 67.5 (d, C-5), 63.3 (d, C-7a), 62.1
Eur. J. Org. Chem. 2013, 4047–4056
© 2013 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim
www.eurjoc.org
4055

J. A. Tamayo, A. Goti et al.
FULL PAPER
[6] a) F. Cardona, C. Parmeggiani, E. Faggi, C. Bonaccini, P. Grat-
teri, L. Sim, T. M. Gloster, S. Roberts, G. J. Davies, D. R. Rose,
A. Goti, Chem. Eur. J. 2009, 15, 1627–1636; b) G. D’Adamio,
A. Goti, C. Parmeggiani, E. Moreno-Clavijo, I. Robina, F.
Cardona, Eur. J. Org. Chem. 2011, 7155–7162; c) F. Cardona,
G. Moreno, F. Guarna, P. Vogel, C. Schuetz, P. Merino, A.
Goti, J. Org. Chem. 2005, 70, 6552–6555; d) I. Delso, T. Tejero,
(t, CH2OH), 35.2 (t, C-7), 16.5 (q, Me) ppm. HRMS (NALDI-
TOF): calcd. for C9H18NO4 [M + H]+ 204.1236; found 204.1237.
Supporting Information (see footnote on the first page of this arti-
cle): Copies of the 1H NMR and 13C NMR spectra of all new
compounds, the synthesis of compounds 9, and tetraacetates 1a
and 2a of indolizidines 1 and 2, respectively, GLC analyses of enzy-
matic acetylation, and glycosidase inhibition assays.
A. Goti, P. Merino, Tetrahedron 2010, 66, 1220–1227; e) ref.[4e]
,
and references cited therein.
[7] a) J. A. Tamayo, F. Franco, F. Sánchez-Cantalejo, Eur. J. Org.
Chem. 2011, 7182–7188; b) J. A. Tamayo, F. Franco, F.
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Acknowledgments
The authors thank the Ministero dell’Università e della Ricerca
(MIUR) (PRIN 2010-11, 2010L9SH3K_006) and Ente CRF for
financial support. The authors also thank the Spanish Ministerio
de Ciencia e Innovación (MICINN) (CTQ2008-01565/BQU) and
Ministerio de Economia y Competitividad (MEC) (CTQ2012-
31247) and the Junta de Andalucía (FQM 345) for financial sup-
port. The Erasmus programme is acknowledged for a mobility
grant to D. M. Maurizio Passaponti and Brunella Innocenti (Uni-
versity of Firenze) are acknowledged for technical support. The
authors thank Martina Bonciani for parts of the synthetic work.
[9] For other recent syntheses of hyacinthacines B, see: a) T. Sen-
goku, T. Satoh, M. Ashima, M. Takahashi, H. Yoda, Tetrahe-
dron 2008, 64, 8052–8058; b) P. V. Reddy, P. Koos, A. Veyron,
A. E. Greene, P. Delair, Synlett 2009, 7, 1141–1143; c) ref.[4g]
;
d) see ref.[5c]
.
[10] For different synthetic strategies affording 5-methylated pyrrol-
izidines and indolizidines, see: a) C. Ribes, E. Falomir, M.
Carda, J. A. Marco, Tetrahedron 2009, 65, 6965–6971; b) X.-G.
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W.-J. Liu, J.-L. Ye, P.-Q. Huang, Org. Biomol. Chem. 2010, 8,
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G. W. J. Fleet, N. Asano, Carbohydr. Res. 1999, 316, 95–103; b)
N. Asano, H. Kuroi, K. Ikeda, H. Kizu, Y. Kameda, A. Kato,
I. Adachi, A. A. Watson, R. J. Nash, G. W. J. Fleet, Tetrahe-
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Molyneux, G. W. J. Fleet, Tetrahedron: Asymmetry 2000, 11,
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Received: January 18, 2013
Published Online: May 6, 2013
4056
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Eur. J. Org. Chem. 2013, 4047–4056