Synthesis and evaluation of dimeric lipophilic iminosugars as inhibitors of glucosylceramide metabolism

Article abstract of DOI:10.1016/j.tetasy.2009.02.043

Four dimeric and four monomeric lipophilic iminosugars were synthesized and subsequently evaluated on their inhibitory potential towards mammalian glucosylceramide synthase, glucocerebrosidase, β-glucosidase 2, sucrase and lysosomal α-glucosidase. Compared to their monomeric counterparts the dimeric inhibitors showed decreased inhibition of glucosylceramide synthase and generally a comparable inhibitory potency for the glycosidases.

Full text of DOI:10.1016/j.tetasy.2009.02.043

Tetrahedron: Asymmetry 20 (2009) 836–846
Contents lists available at ScienceDirect
Tetrahedron: Asymmetry
journal homepage: www.elsevier.com/locate/tetasy
Synthesis and evaluation of dimeric lipophilic iminosugars as inhibitors
of glucosylceramide metabolism
Tom Wennekes a, Richard J. B. H. N. van den Berg a, Kimberly M. Bonger a, Wilma E. Donker-Koopman b,
Amar Ghisaidoobe a, Gijsbert A. van der Marel a, Anneke Strijland b, Johannes M. F. G. Aerts b,
,
*
Herman S. Overkleeft a,
*
a Leiden Institute of Chemistry, Gorlaeus Laboratories, Leiden University, PO Box 9502, 2300 RA Leiden, The Netherlands
b Department of Medical Biochemistry, Academic Medical Center, University of Amsterdam, 1105 AZ, Amsterdam, The Netherlands
a r t i c l e i n f o
a b s t r a c t
Article history:
Four dimeric and four monomeric lipophilic iminosugars were synthesized and subsequently evaluated
Received 2 February 2009
Accepted 24 February 2009
Available online 22 April 2009
on their inhibitory potential towards mammalian glucosylceramide synthase, glucocerebrosidase, b-glu-
cosidase 2, sucrase and lysosomal a-glucosidase. Compared to their monomeric counterparts the dimeric
inhibitors showed decreased inhibition of glucosylceramide synthase and generally a comparable inhib-
itory potency for the glycosidases.
Special edition of Tetrahedron: Asymmetry
in honour of Professor George Fleet’s 65th
birthday
Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction
GBA1
O
Lysosomal
degradation
Glucosylceramide 1 (Fig. 1) and its more complex glycosylated
derivatives are called glycosphingolipids (GSLs). They are compo-
nents of the outer cellular membrane and are involved in many
(patho) physiological processes in man, such as intercellular recog-
nition, signalling processes (e.g., insulin signalling) and interac-
tions with pathogens.1,2 Glucosylceramide 1 functions as the
crucial precursor for the biosynthesis of most complex GSLs. The
biosynthesis of 1 takes place at the cytosolic side of the Golgi appa-
ratus and is catalyzed by the membrane-bound enzyme, glucosyl-
ceramide synthase (GCS). This glycosyltransferase catalyzes the
Biosynthesis
at cytosolic side
Golgi apparatus
HO
HO
HO
( )10
( )10
GCS
HN
O
O
OH
OH
1
Degradation at
plasma membrane
GBA2
Figure 1. Structure of glucosylceramide 1 and an overview of its metabolism.
glycosylation of a-UDP-D-glucopyranose with ceramide to produce
1. The main catabolic pathway of 1 occurs in the lysosome where
glucocerebrosidase (GBA1) with assistance of activator protein
saposin C catalyzes the hydrolysis of the b-glycosidic bond that
connects the ceramide and glucose residues in 1. The membrane-
bound b-glucosidase 2 (GBA2), located on the outside of the plas-
ma membrane can also hydrolyze 1.3,4
In an ongoing study we aim to develop selective inhibitors for
each of the three enzymes, GCS, GBA1 and GBA2.5–8 Selective
inhibitors can be used as tools to further investigate the diverse
functions of GSLs, but also have potential as therapeutics for dis-
eases associated with abnormal GSL metabolism such as lysosomal
sphingolipidoses9,10 and type 2 diabetes.5 We have already shown
that inhibition of GCS through oral dosage of lipophilic iminosugar
2 (Fig. 2) to ob/ob mice, which is a type II diabetes model, downreg-
ulates glycosphingolipid biosynthesis and restores insulin receptor
sensitivity.5 We selected compound 2, a known potent inhibitor of
all three enzymes, as a lead compound in our research efforts.6,8
OH
N
OH
N
HO
HO
HO
HO
O
O
2
3
OH
OH
2
2
1
1
A
B
* Corresponding authors.
E-mail addresses: j.m.aerts@amc.uva.nl (J.M.F.G. Aerts), h.s.overkleeft@chem.lei-
denuniv.nl (H.S. Overkleeft).
Figure 2. Structure of lead compound 2, its
of the two bis-functionalized adamantanes (A and B).
L-ido derivative 3 and the general design
0957-4166/$ - see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tetasy.2009.02.043

T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
837
The structure of 2 can be divided into three parts, the iminosugar
core, a pentyl spacer and the adamantan-1-yl-methoxy hydropho-
bic moiety. There is literature precedence that C-5-epimerized
or shortening the pentyl spacer decreases inhibitory potency, as
is the case with removal of the ether function. Also, hydrophobic
N-alkylated iminosugars having a ether functionality in the N-alkyl
portion are thought to be less cytotoxic than isosteric compounds
without this functional group.19 The orientation of the two binding
elements in a bivalent inhibitor can also be critical. In this respect
the adamantane moiety, besides targeting the inhibitors to the
membrane, also functions as a rigid scaffold.20 If the adamantane
moiety indeed binds in hydrophobic lipid bilayer pockets then
the two pentyl-spaced iminosugars should be oriented in the same
direction in order to potentiate bivalent action as depicted in Fig-
ure 3. To this end we designed bifunctional adamantane A
(Fig. 2). Scaffold A, however, does introduce an additional methy-
lene in between the adamantane and the iminosugar. Therefore,
a second difunctionalized adamantane B was designed. Scaffold B
minimizes the change to the general design of the original inhibitor
(2 and 3), but at the cost of the orientation of the two iminosugar
moieties.
The research presented here describes the synthesis of the two
types of dimeric scaffolds functionalized with two 1-deoxynojiri-
mycin or L-ido-1-deoxynojirimycin iminosugars. To investigate
the effect of the added methylene in scaffold A the monomeric ana-
logues of this scaffold were also prepared. The resulting six com-
pounds were evaluated together with known 2 and 3 in an
enzyme assay for inhibition of the three glucosylceramide metab-
olism-related enzymes (GBA1, GBA2 and GCS) and two non-related
L
-ido-1-deoxynojirimycin derivatives are viable GCS inhibitors.11,12
Indeed we observed that L-ido derivative 3 (Fig. 2) is a selective GCS
inhibitor with a potency comparable to that of 2.13
Results from our previous studies have also shown the impor-
tance of the adamantane moiety in achieving potent inhibition,
especially for GBA2 and GCS. Both these enzymes are membrane-
bound and the role of the hydrophobic adamantane might be to
concentrate the iminosugar inhibitor at the location of these en-
zymes. If this hypothesis holds true, then appending a second imi-
nosugar to the adamantane may increase the local concentration of
the iminosugar near the enzyme’s active site even further, with an
altered inhibitory potency as a result.
In nature, related bivalent or even multivalent interactions are
often encountered as a way of improving the interactions between
receptor and ligand.14 Multivalency is also adopted as a strategy in
pharmaceutical research to improve interactions of natural recep-
tors with designed ligands.14,15 The most basic form of multiva-
lency is bivalency and in general a bivalent ligand can bind to its
target receptor in four ways that all cause increased specificity
and/or strength of binding (Fig. 3).15
A
B
glycosidases (sucrase and lysosomal a-glucosidase).
2. Results and discussion
Reductive amination was chosen as the means for condensation
of the iminosugar cores to the difunctionalized adamantane scaf-
folds. This entailed the synthesis of dipentanal derivatized ada-
mantanes that in turn are accessible by ozonolysis of dihex-1-
ene precursors. The synthesis of the target compounds starts with
the preparation of the two adamantane cores. Generation of the
enolate of commercially available 1-adamantan-acetic acid 4 and
condensation with formaldehyde produced 5 in 27% yield with
68% recovery of 4 (Scheme 1).21 Reduction of 5 with LiAlH4 pro-
vided 6.
C
D
Upon increasing the scale of the aldol condensation with 4 the
yield of 5 decreased due to the difficulties in handling gaseous
formaldehyde. Therefore, a different route was developed. The
ethyl ester 7 of 4 was prepared and deprotonated with LDA. Reac-
tion of this enolate with ethyl chloroformate produced malonate
derivative 8 (82% yield) that upon reduction with LiAlH4 also gave
6. Reduction of 1,3-adamantan-diacetic acid 9 gave diol 10. Wil-
liamson etherification of 6 and 10 with excess 6-bromo-1-hexene
produced almost no dialkylated product, but instead yielded pre-
dominantly the mono-alkylated products. Successively performing
the deprotonation and bromide addition twice in one-pot did pro-
duce dienes 11 and 12. Ozonolysis of 11 and workup with dimeth-
Figure 3. Overview of mechanisms of bivalent ligand binding. (A) Rapid rebinding
of inhibitor by the high local concentration of ‘free’ ligand; (B) increased binding
specificity and strength by additional binding at an allosteric site; (C) simultaneous
inhibition of two neighbouring copies of the receptor; (D) dimerization/clustering
of the two copies of the receptor.
There is currently no evidence that mammalian glycosyltrans-
ferases or glycosidases homodimerize or possess allosteric binding
sites. Therefore a potential bivalent inhibitor of these enzymes
would operate via mechanism A in Figure 3. Although the multiva-
lency approach has been extensively used in optimization of recep-
tor–ligand interactions it is relatively unexplored in the
development of glycosidase/glycosyltransferase inhibitors. A liter-
ature search revealed only a few examples of previous studies with
iminosugar-based multivalent glycosidase inhibitors.16–18 These
examples differ from the research presented here in that the mul-
timeric inhibitors developed in those studies all target soluble en-
zymes, whereas GCS and GBA2 targeted in our research are
membrane-bound.
ylsulfide produced
a
mixture of products. Isolation and
characterization of the major product (ꢀ30%) showed that it was
not dialdehyde 13 but rather an intermediate ozonide (14 or 15,
see note22). Ozonolysis of 11 and 12 and treatment of the interme-
diate ozonides with trimethylphosphine did provide dialdehydes
13 and 16. Reductive amination of the dialdehydes with either
2,3,4,6-tetra-O-benzyl-1-deoxynojirimycin7 or 2,3,4,6-tetra-O-
benzyl-L
-ido-1-deoxynojirimycin13 provided the four penultimate
compounds 17, 18, 19 and 20. Palladium-catalyzed hydrogenation
at 4 bar produced the dimeric compounds 21, 22, 23 and 24. The
two monomeric reference analogues 27 and 28 were prepared by
selective nitrogen alkylation of 1-deoxynojirimycin and L-ido-1-
deoxynojirimycin with bromide 26.
A critical aspect in the design of the bivalent inhibitors is the
type and distance of separation between the two iminosugars.
For inhibitor 2 it has already been ascertained that lengthening

838
T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
O
O
O
O
HO
OH
OH
HO
EtO
c
9
route A
route B
4
7
a
d
b
e
O
O
O
HO
HO
OH
HO
O
OH
EtO
OEt
b
b
10
8
5
6
e
O
f
O
R
R
R
O
R
11: R = CH2
13: R = O
12: R = CH2
16: R = O
f
g
g
OR
N
RO
OR
N
RO
N
OR
RO
RO
OR
OR
RO
RO
O
O
O
O
N
O
O
O
19
23
17
21
OR
h
h
OR
OR
OR
OR
OR
R = Bn
R =H
h
OR
N
RO
N
OR
N
RO
N
RO
RO
OR
OR
RO
RO
OR
OR
O
18
22
20
24
h
h
j
OR
OR
OR
OH
OH
N
R
O
HO
HO
HO
HO
j
N
O
O
26
26
25: R = OH
26: R = Br
i
27
28
OH
OH
Scheme 1. Synthesis of dimeric lipophilic iminosugars 21–24 and reference monomeric analogues 27 and 28. Reagents and conditions: (a) (i) LDA (2 equiv) THF, ꢁ30 °C, 1 h;
(ii) HMPA (1 equiv), ꢁ30 °C; (iii) formaldehyde gas, ꢁ30 °C, 20 min 3 mmol scale: 4: 68% 5: 27%; 39 mmol scale: 4: 88% 5: 11%; (b) LiAlH4, THF, 0 °C?rt, 20 h, 6 from 5: 86%; 6
from 8: 96%; 10: used crude; (c) EtOH, 18 M H2SO4, reflux, 5 h, 93%. (d) (i) LDA (1 equiv) THF, ꢁ30 °C, 1 h; (ii) HMPA (1 equiv), ꢁ30 °C; (iii) ethyl chloroformate, ꢁ30 °C, 1 h,
82%; (e) (i) 6-bromo-1-hexene, NaH, TBAI, DMF, 0 °C to rt, 2 h; (ii) 6-bromo-1-hexene, NaH, 20 h, 11: 72%; 12: 65%; (f) (i) O3, DCM, ꢁ30 °C, 30 min; (ii) PMe3, rt, 4 h, 13: 80%;
16: 72%; (g) 2,3,4,6-tetra-O-benzyl-1-deoxynojirimycin or 2,3,4,6-tetra-O-benzyl-L-ido-1-deoxynojirimycin (3 equiv), NaCNBH3 (6 equiv), Na2SO4 (5 equiv), EtOH/AcOH (20/
1), 20 h, 0 °C?rt, 17: 60%; 18: 55%; 19: 62%; 20: 64%; (h) Pd/C, H2 4 bar, n-propanol/EtOAc, HCl, 20 h, 21: 79%; 22: 73%; 23: 60%, 24: 55%; (i) CBr4, PPh3, CH3CN, reflux, 3 h,
87%; (j) 1-deoxynojirimycin or -ido-1-deoxynojirimycin (0.66 equiv), K2CO3 (2 equiv), DMF, 90 °C, 48 h, 27: 58%; 28: 43%.
L
2.1. Biological evaluation
upon elution (25% MeOH and 5% NH4OH in CHCl3) from a silica gel
column as opposed to almost complete retention for 21 (Rf =
0–0.05). The increased polarity of the dimeric compounds might
negatively affect their cell permeability as well as their concentra-
tion in the Golgi membranes where GCS resides. The cell permeabil-
ity of compounds 21–24 was investigated by testing them in an
in vitro assay for GCS inhibition. All four compounds showed compa-
rable IC50 values to the in vivo assay indicating that cell permeability
is probably not the cause for the decreased GCS inhibition.
Although GCS inhibition decreases for all compounds tested
when compared with 2 and 3, they are still more or equally potent
as the commercial GCS inhibitor and Gaucher therapeutic,23 N-bu-
tyl-1-deoxynojirimycin (Miglustat, Zavesca). This compound,
which was first synthesized in 1988,23 is currently used as a ther-
apeutic for Gaucher disease.24 A trend observed in the GCS inhibi-
The six compounds were evaluated and compared to 2 and 3 in
an enzyme assay for inhibition of the three glucosylceramide
metabolism-related enzymes, GCS, GBA1 and GBA2 (Table 1).
Introduction of an additional methylene between the ether and
adamantane as in 27 and 28 hardly affects the inhibition profile
when compared to 2 and 3. The dimeric derivatives 21 and 22 also
inhibit GBA1 and GBA2 comparable to the monomeric 27 and 28,
but are markedly less potent for GCS. The second type of dimeric
iminosugars 23 and 24 again showed little change in inhibitory po-
tency for GBA1 and GBA2. However, also for this type of dimeric
iminosugars the inhibition of GCS decreases considerably upon
introduction of a second pentyl-spaced iminosugar moiety.
Besides the steric implications of attaching a second iminosugar-
pentyl moiety to the adamantane there is also a marked effect upon
the overall polarity of the molecule: the number of hydroxyls are
doubled. This phenomenon was already observed during purifica-
tion of the bivalent end products. Lead compound 2 has an Rf of 0.3
tory data is that the
inhibitors of GCS than their
L
-ido analogues are more potent and selective
-gluco equivalents. This corresponds
D
with our recent identification of 3 as a more potent and selective
inhibitor of GCS than 2.13

Table 1
Enzyme inhibition assay results (GCS: % inhibition at
l
M; the four glycosidases: apparent IC50 values in
GCS in vivo
l
M)
GCS in vitro
Compound
GBA1 in vitro
GBA2 in vitro
Lysosomal
a-glucosidase in vitro
Sucrase in vitro
(%)a
(lM)
(%)b
(lM)
OH
HO
N
O
O
2: C-5 = (R)
3: C-5 = (S)
D
-gluco
50
75
0.2
0.1
36
—
0.5
—
0.2
2
0.001
0.03
0.4
>100
0.5
>100
L-ido
HO
OH
OH
N
HO
HO
27: C-5 = (R)
28: C-5 = (S)
D-gluco
55
43
1
1
40
68
5
5
0.19
2
0.005
0.006
0.8
>100
0.35
200
L
-ido
OH
OH
N
HO
5
N
5
O
HO
HO
OH
OH
O
21: C-5 = (R)
22: C-5 = (S)
D
-gluco
30
69
20
20
64
52
40
10
0.38
5
0.008
0.150
0.35
700
0.4
200
L
-ido
OH
OH
OH
N
HO
5
N
5
HO
HO
OH
OH
O
O
23: C-5 = (R)
24: C-5 = (S)
D
-gluco
35
46
20
5
47
59
40
10
0.34
6
0.010
0.013
0.5
>100
0.5
>100
L
-ido
OH
OH
a
Average of three measurements.
One measurement.
b

840
T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
GBA2 and GCS are both membrane-bound enzymes that to-
for 7 days. Rf values were determined from TLC analysis using
DC-fertigfolien (Schleicher & Schuell, F1500, LS254) with detection
by spraying with a solution of (NH4)6Mo7O24 ꢂ 4H2O (25 g/L) and
(NH4)4Ce(SO4)4 ꢂ 2H2O (10 g/L) in 10% sulfuric acid or a solution
of phosphomolybdic acid hydrate (7.5 wt % in ethanol) followed
by charring at ꢀ150 °C. Visualization of all deprotected iminosugar
compounds during TLC analysis was accomplished by exposure to
iodine vapour. Column chromatography was performed on silica
gether with GBA1 process membrane-localized lipophilic sub-
strates. To further test the bivalent properties of 21–24 they
were also evaluated as inhibitors of the non-membrane-bound
lysosomal
crase that both process non-lipophilic substrates. Both are potently
inhibited by lead compound 2, but not by -ido-analogue 3. The re-
sults of the enzyme assay confirm the fact that iminosugar-type
inhibitors of lysosomal -glucosidase and sucrase require -glu-
a-glucosidase and the intestinal membrane-bound su-
L
a
D
gel (40–63 l
m). 1H and 13C-APT NMR spectra were recorded on a
cose stereochemistry. Both bivalent iminosugars 21 and 23 show
similar inhibition of the enzymes and are comparable to lead com-
pound 2. Bivalent 21 is twice as potent as its monomeric derivative
Bruker DMX 600 (600/150 MHz), Bruker DMX 500 (500/
125 MHz) or Bruker AV 400 (400/100 MHz) spectrometer in CDCl3
or MeOD. Chemical shifts are given in ppm (d) relative to tetra-
methylsilane as internal standard (1H NMR in CDCl3) or the signal
of the deuterated solvent. Coupling constants (J) are given in hertz.
Where indicated, NMR peak assignments were made using COSY
and HSQC experiments. All presented 13C-APT spectra are proton
decoupled. High resolution mass spectra were recorded by direct
27 for lysosomal
a-glucosidase but this trend is not observed for
inhibition of sucrase.
3. Conclusion
injection (2 lL of a 2 lM solution in water/acetonitrile; 50/50; v/
The synthesis and evaluation of four monomeric and four di-
meric lipophilic iminosugars 21–24 are reported. Evaluation of
two novel monomeric analogues 27 and 28 and two known lipo-
philic iminosugars 2 and 3 showed that the structure–activity rela-
tionship (SAR) of GBA1, GBA2 and GCS inhibition tolerates the
introduction of an additional methylene between the ether func-
tion and the adamantane. The four dimeric iminosugars 21–24 in-
hibit GBA1 and GBA2 with a potency comparable to that of their
monomeric counterparts 2, 3, 27 and 28. However, inhibition of
GCS decreased markedly for all dimeric compounds. This was ob-
served in both in vivo and in vitro GCS assays and thus reduced cell
permeability of the more polar dimeric iminosugars was ruled out
as a cause for the decrease.
v and 0.1% formic acid) on a mass spectrometer (Thermo Finnigan
LTQ Orbitrap) equipped with an electrospray ion source in positive
mode (source voltage 3.5 kV, sheath gas flow 10, capillary temper-
ature 250 °C) with resolution R = 60,000 at m/z 400 (mass range m/
z = 150–2000) and dioctylphthalate (m/z = 391.28428) as a ‘lock
mass’. The high resolution mass spectrometer was calibrated prior
to measurements with a calibration mixture (Thermo Finnigan).
Optical rotations were measured on a Propol automatic polarime-
ter (Sodium D-line, k = 589 nm). ATR-IR spectra were recorded on a
Shimadzu FTIR-8300 fitted with a single bounce Durasample IR
diamond crystal ATR-element and are reported in cmꢁ1
.
Enzyme assays: Lipids were extracted according to Folch et al.29
Neutral GSLs were analyzed as described previously.30 Ganglioside
composition was determined as described previously.31 IC50 values
of the iminosugars for the various enzyme activities were deter-
mined by exposing cells or enzyme preparations to an appropriate
range of iminosugar concentrations (DMSO stock solutions diluted
with RPMI medium). In vivo glucosylceramide synthase assay: the
mouse macrophage cell line RAW-267 was grown to 90–100% con-
fluence in growth medium at 37 °C in a 5% CO2 incubator. The
growth medium consisted of RPMI-1640 + 10% FCS + penicillin
Our results seem to indicate that dimeric lipophilic iminosugars
do not inhibit GBA1, GBA2, GCS or lysosomal a-glucosidase and su-
crase in a bivalent fashion (as in mode A in Fig. 3). However, the
retained inhibitory potency for GBA1, GBA2 and GCS does show
that the SAR tolerates the attachment of a second pentyl-spaced
iminosugar, with no clear difference between the two adamantane
scaffolds. It is possible that the length of the pentyl spacer is
impeding bivalent binding modes, and therefore variation of its
length is worth investigating in future analogues. The much-in-
creased polarity of the presented bivalent inhibitors might make
them less lipophilic and thereby less available for the mem-
brane-bound enzymes. Targeting to the membrane-bound en-
zymes and stabilization inside the membrane could therefore
perhaps be improved in future dimeric analogues of 2 and 3 by
basing the scaffold on the more hydrophobic diamondoids,
diamantane and triamantane.25–28
(150 lg/mL) and streptomycin (250 lg/mL) + 50 mM Hepes buffer
(4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid). The incuba-
tion flask was washed with RPMI medium without serum
(3 ꢂ 5 mL) to remove serum. Cells were taken up in 3 mL
RPMI + 50 mM Hepes + 300 lM conduritol B epoxide (10 mM stock
in RPMI) + the inhibitor (0.1, 1, 10
diluted in RPMI) and 5 nmol C6-NBD-ceramide/BSA complex
(N-[7-(4-nitrobenzo-2-oxa-1,3-diazole)]-6-aminocaproyl- -erythro-
lM (20 mM stock in DMSO
D
sphingosine) were added successively to the cell culture. The cells
were incubated for 1 h at 37 °C in a 5% CO2 incubator. At the end of
incubation, the flask was inspected for potential cytotoxicity of the
inhibitor and washed with medium without serum (5 ꢂ 5 mL). A
50 mM potassium phosphate buffer (KPi buffer, pH 5.8, 0.75 mL)
was added and the flask was placed in ice. Cells were removed
from the flask by scraping and the harvested cells were collected
in a capped plastic vial and immediately immersed in liquid nitro-
gen. The frozen cell lysate (ꢀ0.75 mL) was suspended in methanol
(3 mL) and extracted with chloroform (3 mL). After extraction a
0.73% NaCl solution (2.0 mL) was added to the biphasic. The aque-
ous phase was extracted once more with chloroform (1 mL). The
combined chloroform layers were isolated and concentrated at
30–40 °C under a nitrogen flow. Lipids were separated by thin-
layer chromatography (HP-TLC plates 20 ꢂ 10 Silica Gel 60 van
Merck) using chloroform/methanol/15 mM aq CaCl2 (60:35:8, v/
v/v) as the developing solvent. The C6-NBD-labelled (glyco)sphin-
golipids were identified using standards, visualized with a Ty-
phoon Trio Variable Mode Imager (kex 488 nM, kem 520 nM) and
4. Experimental
4.1. General methods
All solvents and reagents were obtained commercially and used
as received unless stated otherwise. Reactions were executed at
ambient temperatures unless stated otherwise. All moisture sensi-
tive reactions were performed under an argon atmosphere. Resid-
ual water was removed from starting compounds by repeated
coevaporation with dioxane, toluene or dichloroethane. All sol-
vents were removed by evaporation under reduced pressure. Reac-
tion grade acetonitrile, n-propanol and methanol were stored on
3 Å molecular sieves. Other reaction grade solvents were stored
on 4 Å molecular sieves. THF was distilled prior to use from LiAlH4.
Ethanol was purged of acetaldehyde contamination by distillation
from zinc/KOH. DCM was distilled prior to use from P2O5. Diisopro-
pylamine was distilled from KOH and stored over KOH. Parafor-
maldehyde was dried prior to use in a P2O5 containing desiccator

T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
841
quantified with IMAGEQUANT TL software. Glucocerebrosidase activity
was measured using recombinant enzyme and 4-methylumbellife-
ryl-b-glucose as substrate.32 GBA2 was measured using enzyme-
containing membrane preparations from Gaucher spleen and 4-
2), 1.95 (s, 3H, 3 ꢂ CH Ada), 1.88–1.45 (m, 12H, 6 ꢂ CH2 Ada). 13C
NMR (50 MHz, MeOD) d 177.6 (C(O)-1), 61.5 (CH-2), 60.6 (CH2-
3), 41.7 (CH2 Ada), 38.1 (CH2 Ada), 35.0 (Cq Ada), 30.2 (CH Ada).
IR
m
max(thin film)/cmꢁ1: 3332, 2902, 2851, 1698, 1445, 1257,
methylumbelliferyl-b-glucoside as substrate.8 Lysosomal
a
-gluco-
sidase was measured using purified enzyme from human urine
and 4-methylumbelliferyl-
-glucoside as substrate.8 Sucrase activ-
1223, 1008, 808, 652, 618. HRMS: found 225.1485 [M+H]+, calcu-
lated for [C13H20O3 + H]+ 225.1485.
a
ity was determined with homogenates of freshly isolated rat intes-
tine by measuring liberated glucose from the corresponding
disaccharides.33 In vitro assay of GCS activity was conducted with
in spleen microsomes. Compounds 21–24 were tested as their
dihydrochloric acid salt and 27 and 28 were assayed as their
TFA-salt.
General procedure A—Reductive amination of dialdehydes 13 and
16: A dry and cooled (0 °C) mixture of the dialdehyde (1 equiv),
Na2SO4 (5 equiv) and 2,3,4,6-tetra-O-benzyl-1-deoxynojirimycin
(3 equiv; synthesis as reported previously6,7) or 2,3,4,6-tetra-O-
4.1.2. 2-(Adamantan-1-yl)propane-1,3-diol 6
Reduction of 5: A dry and cooled (0 °C) solution of 5 (424 mg,
2.0 mmol) in THF (20 mL) was charged with LiAlH4 (5 mL,
5.0 mmol; 1 M in THF), stirred for 20 h and warmed to rt. The reac-
tion was quenched (1st EtOAc 30 min, 2nd water) and aq 1 M HCl
(30 mL) was added. The mixture was extracted with EtOAc
(2 ꢂ 50 mL) and the resulting combined organic phase was washed
with sat aq NaHCO3 (100 mL). The organic phase was dried
(Na2SO4) and concentrated. The resulting residue was purified by
silica gel column purification (2:1 EtOAc/PE?4:1 EtOAc/MeOH)
to furnish 6 (339 mg, 1.73 mmol) as white solid in 86% yield.
Reduction of 8: Compound 8 (1.07 g, 3.64 mmol) was subjected to
same procedure as described above, but with 4.1 equiv of LiALH4
to produce 6 (685 mg, 3.49 mmol) in 96% yield after purification.
Rf = 0.52 (19:1; EtOAc/AcOH). 1H NMR (200 MHz, MeOD) d 3.84
(dd, J = 4.0, 10.7, 2H, CHH-1,3), 3.66 (dd, J = 7.3, 10.8, 2H, CHH-
1,3), 1.94 (s, 3H, 3 ꢂ CH Ada), 1.84–1.59 (m, 12H, 6 ꢂ CH2 Ada),
1.26–1.12 (m, 1H, CH-2). 13C NMR (50 MHz, MeOD) d 61.5 (CH2-
1,3), 54.2 (CH-2), 41.7 (CH2 Ada), 38.4 (CH2 Ada), 35.2 (Cq Ada),
benzyl-L-ido-1-deoxynojirimycin (3 equiv; synthesis as reported
previously13) in EtOH/AcOH (20/1, v/v, 0.1 M) was charged with
NaCNBH3 (6 equiv). The reaction mixture was stirred for 20 h and
allowed to warm to rt. The reaction mixture was diluted with
EtOAc (100 mL) and washed with sat aq NaHCO3 (2 ꢂ 100 mL).
The organic phase was dried (Na2SO4) and concentrated. The crude
product was purified by silica gel column purification (1:5 EtOAc/
PE?1:1 EtOAc/PE) to afford the product. TLC-analysis: Rf 13 = 0.55;
16 = 0.60 (2:3; EtOAc/PE).
General procedure B—Pd/C-catalyzed hydrogenolysis: A solution of
30.3 (CH Ada). IR m
max(thin film)/cmꢁ1: 3235, 2900, 2846, 1447,
compound (ꢀ300–400
lmol) in n-propanol/EtOAc (50 mL, 8/1, v/v)
1344, 1034, 1009, 973. HRMS: found 211.1694 [M+H]+, calculated
for [C13H22O2 + H]+ 211.1693.
was acidified to pH ꢀ2 with 1 M aq HCl (1 mL). Argon was passed
through the solution for 5 min, after which a catalytic amount of
Pd/C (50 mg, 10 wt % on act. carbon) and Pd black (5 mg) was
added. The reaction vessel was placed under vacuum and subse-
quently ventilated with hydrogen gas. This cycle was repeated
one more time after which the vessel was placed under 4 bar of
hydrogen gas and mechanically shaken for 20 h. Pd/C was removed
by filtration over a glass microfibre filter, followed by thorough
rinsing of the filter cake with MeOH. The filtrate was concentrated
with coevaporation of toluene. The residue was purified by silica
gel column purification (3:1 EtOAc/MeOH+5%NH4OH?19:1
MeOH/NH4OH) to give the product. Silica gel residue in the puri-
fied product was removed by dissolving the product in MeOH/
4.1.3. Ethyl (adamantan-1-yl)acetate 7
Sulfuric acid (1 mL, 98%/18 M) was added to a solution of 1-ada-
mantan-acetic acid 4 (10 g, 51.5 mmol) in ethanol (200 mL). The
reaction mixture was refluxed for 5 h after which it was neutral-
ized by addition of 4 M aq NaOH (ꢀ4.5 mL). The reaction mixture
was concentrated to a quarter of its volume and EtOAc (100 mL)
was added. The mixture was washed with sat aq NaHCO3
(4 ꢂ 50 mL). The organic phase was dried (Na2SO4) and concen-
trated. The resulting residue was purified by silica gel column puri-
fication (100% PE?1:9 EtOAc/PE) to furnish 7 (2.07 g, 9.3 mmol) as
a colourless oil in 93% yield. Rf = 0.65 (1:12.5; EtOAc/PE). 1H NMR
(400 MHz, CDCl3) d 4.11 (q, J = 7.1, 2H, CH2 ethyl), 2.05 (s, 2H,
CH2-COOEt), 1.97 (s, 3H, 3 ꢂ CH Ada), 1.72–1.62 (m, 12H, 6 ꢂ CH2
Ada), 1.26 (t, J = 7.1, 3H, CH3 ethyl). 13C NMR (100 MHz, CDCl3) d
171.8 (C@O), 59.9 (CH2 Et), 49.1 (CH2-COOEt), 42.5 (CH2 Ada),
36.9 (CH2 Ada), 35.9 (Cq Ada), 28.7 (CH Ada), 14.5 (CH3 Et). IR
CHCl3 (1/9, v/v), passing the solution over a filter (0.5
centrating the filtrate.
lm) and con-
4.1.1. (R/S)-2-(Adamantan-1-yl)-3-hydroxypropanoic acid 5
Butyllithium (4.25 mL, 6.8 mmol; 1.6 M in hexane) was added
to
a
dry and cooled (ꢁ30 °C) solution of diisopropylamine
m
max(thin film)/cmꢁ1: 2901, 2848, 1731, 1451, 1323, 1255, 1196,
(0.95 mL, 6.8 mmol) in THF (20 mL). After 10 min, a dry solution
of 1-adamantan-acetic acid (4: 602 mg, 3.1 mmol) in THF (15 mL)
was added over a 1 min period at ꢁ30 °C, a yellow/orange turbid
reaction mixture was formed. After 20 min, HMPA (0.56 mL,
3.1 mmol) was added that resulted in a clear orange reaction mix-
ture. Paraformaldehyde (3–4 g) was decomposed at 200 °C and the
resulting formaldehyde vapours were passed by a N2 flow over the
surface of the cooled (ꢁ30 °C) reaction mixture. After complete
depolymerization of the paraformaldehyde the reaction mixture
was stirred for an additional 30 min. The reaction mixture was
quenched by addition of water (1 mL). Ethylacetate (50 mL) was
added and the mixture was extracted with aq 0.1 M HCl
(4 ꢂ 50 mL). The organic phase was dried (Na2SO4) and concen-
trated. The resulting residue was purified by silica gel column puri-
fication (2:1 EtOAc/PE?19:1 EtOAc/AcOH) to produce 5 (185 mg,
0.82 mmol) as white solid in 27% yield with 68% recovery of 1-ada-
mantaneacetic acid. Rf = 0.65 (19:1; EtOAc/AcOH); Rf 1-adaman-
taneacetic acid = 0.85 (19:1; EtOAc/AcOH). 1H NMR (200 MHz,
MeOD) d 3.92–3.71 (m, 2H, CH2-3), 2.23 (dd, J = 4.5, 9.9, 1H, CH-
1135, 1033, 700. HRMS: found 222.1694 [M+H]+, calculated for
[C14H22O2 + H]+ 222.1693.
4.1.4. Diethyl 2-(adamantan-1-yl)malonate 8
Butyllithium (3.75 mL, 6.0 mmol; 1.6 M in hexane) was added
to
a
dry and cooled (ꢁ30 °C) solution of diisopropylamine
(0.84 mL, 6.0 mmol) in THF (10 mL). After 10 min, a dry solution
of 7 (1.20 g, 5.4 mmol) in THF (10 mL) was added over a 1 min per-
iod at ꢁ50 °C, a yellow turbid reaction mixture was formed. After
20 min, HMPA (0.95 mL, 5.4 mmol) was added that resulted in a
clear yellow reaction mixture. Ethyl chloroformate (0.62 mL,
6.4 mmol) was added and the reaction mixture was stirred at
(ꢁ30 °C for 1 h. The reaction mixture was quenched with water
and EtOAc (100 mL) was added. The mixture was extracted with
aq 0.1M HCl (4 ꢂ 50 mL). The organic phase was dried (Na2SO4)
and concentrated. The resulting residue was purified by silica gel
column purification (1:19 EtOAc/PE?1:9 EtOAc/PE) to produce 8
(1.30 g, 4.41 mmol) as a colourless oil in 82% yield. Rf = 0.45
(1:12.5; EtOAc/PE). 1H NMR (400 MHz, CDCl3) d 4.18 (q, J = 7.1,

842
T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
4H, 2 ꢂ CH2 Et), 3.08 (s, 1H, CH-2), 1.99 (s, 3H, 3 ꢂ CH Ada), 1.79 (d,
J = 2.7, 6H, 3 ꢂ CH2 Ada), 1.72–1.62 (m, 6H, 3 ꢂ CH2 Ada), 1.27 (t,
J = 7.1, 6H, 2 ꢂ CH3 Et). 13C NMR (100 MHz, CDCl3) d 168.0 (C(O)-
1,3), 62.7 (CH-2), 60.9 (CH2 Et), 40.0 (CH2 Ada), 36.9 (CH2 Ada),
added. After stirring for 15 min, additional 6-bromo-1-hexene
(0.4 mL) was added and the reaction mixture was stirred for
20 h, warming to rt. The reaction mixture was quenched with
water. The mixture was diluted with Et2O (100 mL) and washed
with water (3 ꢂ 100 mL). The organic phase was dried (Na2SO4)
and concentrated. The resulting residue was purified by silica gel
column purification (100% PE?1:9 EtOAc/PE) to produce 12
(233 mg, 0.65 mmol) as a colourless oil in 65% yield. Rf = 0.90
(1:4; EtOAc/PE). 1H NMR (600 MHz, CDCl3) d 5.81 (ddt, J = 6.7,
10.2, 17.1, 2H, 2 ꢂ @CH-5 hexenyl), 5.00 (d, J = 17.2, 2H,
2 ꢂ @CHH-6 hexenyl), 4.94 (d, J = 10.3, 2H, 2 ꢂ @CHH-6 hexenyl),
3.39–3.35 (m, 4H, 2 ꢂ CH2-1 hexenyl), 2.99 (s, 4H, 2 ꢂ OCH2-
Ada), 2.11–2.05 (m, 4H, 2 ꢂ CH2-4 hexenyl), 2.03 (s, 2H, CH-5,7
Ada), 1.65–1.60 (m, 2H, CH2-6 Ada), 1.60–1.53 (m, 4H, 2 ꢂ CH2-3
hexenyl), 1.46 (ddd, J = 9.4, 17.9, 23.5, 12H, CH2-4,8,9,10 Ada,
2 ꢂ CH2-2 hexenyl), 1.31 (s, 2H, CH2-2 Ada). 13C NMR (150 MHz,
CDCl3) d 139.1 (@CH-5 hexenyl), 114.6 (@CH2-6 hexenyl), 81.8
(OCH2-Ada), 71.6 (CH2-1 hexenyl), 42.0 (CH2-2 Ada), 39.6
(CH2-4,8,9,10 Ada), 37.0 (CH2-6 Ada), 34.7 (Cq-1,3 Ada), 33.8
(CH2-4 hexenyl), 29.2 (CH2-2 hexenyl), 28.5 (CH-5,7 Ada), 25.7
36.1 (Cq Ada), 28.7 (CH Ada), 14.4 (CH3 Et). IR
mmax(thin film)/
cmꢁ1: 2904, 2850, 1753, 1726, 1449, 1368, 1319, 1250, 1221,
1201, 1142, 1032. MS (ESI): found 295.3 [M+H]+, calculated for
[C17H26O4 + H]+ 295.2.
4.1.5. 1,10-[2-(Adamantan-1-yl)propane-1,3-diyl]bis(oxy)dihex-
5-ene 11
A dry cooled (0 °C) solution of 6 (210 mg, mmol) and TBAI
(50 mg, 0.14 mmol) in DMF (5 mL) was charged with NaH
(120 mg, 3 mmol; 60% in mineral oil). The reaction mixture was
stirred for 1 h at after which 6-bromo-1-hexene (0.4 mL, 3 mmol)
was added. The reaction mixture was stirred for 2 h and warmed
to rt. The reaction mixture was cooled and additional NaH
(120 mg) was added. After stirring for 15 min, additional 6-bro-
mo-1-hexene (0.4 mL) was added and the reaction mixture was
stirred for 20 h, warming to rt. The reaction mixture was quenched
with water. The mixture was diluted with Et2O (100 mL) and
washed with water (3 ꢂ 100 mL). The organic phase was dried
(Na2SO4) and concentrated. The resulting residue was purified by
silica gel column purification (100% PE?1:9 EtOAc/PE) to produce
11 (270 mg, 0.72 mmol) as a colourless oil in 72% yield. Rf prod-
uct = 0.85; mono-alkylated side product = 0.38 (1:5; EtOAc/PE).
1H NMR (600 MHz, CDCl3) d 5.81 (ddt, J = 6.7, 10.2, 17.0, 2H,
2 ꢂ @CH-5 hexenyl), 5.04–4.98 (m, 2H, 2 ꢂ @CHH-6 hexenyl),
4.97–4.91 (m, 2H, 2 ꢂ @CHH-6 hexenyl), 3.52 (dd, J = 4.2, 9.4, 2H,
2 ꢂ CHH-1,3 propyl), 3.45–3.33 (m, 6H, 2 ꢂ CHH-1,3 propyl,
2 ꢂ CH2-1 hexenyl), 2.07 (dd, J = 7.2, 14.5, 4H, 2 ꢂ CH2-4 hexenyl),
1.93 (s, 3H, 3 ꢂ CH Ada), 1.71–1.60 (m, 12H, 6ꢂCH2 Ada), 1.60–1.54
(m, 4H, 2 ꢂ CH2-2 hexenyl), 1.49–1.42 (m, 4H, 2 ꢂ CH2-3 hexenyl),
1.32–1.26 (m, 1H, CH-2 propyl). 13C NMR (150 MHz, CDCl3) d 139.1
(@CH-5 hexenyl), 114.6 (@CH2-6 hexenyl), 70.9 (CH2-1 hexenyl),
68.3 (CH2-1,3 propyl), 49.5 (CH-2 propyl), 40.8 (CH2 Ada), 37.4
(CH2 Ada), 34.1 (Cq Ada), 33.7 (CH2-4 hexenyl), 29.4 (CH2-2 hexe-
(CH2-3 hexenyl). IR
m : 2899, 2848, 1640,
max(thin film)/cmꢁ1
1455, 1366, 1108, 993, 908. HRMS: found 361.3102 [M+H]+, calcu-
lated for [C24H40O2 + H]+ 361.3101.
4.1.7. 5,50-[2-(Adamantan-1-yl)propane-1,3-diyl]bis(oxy)-
dipentanal 13
A solution of 11 (400 mg, 1.1 mmol) in DCM (70 mL; EtOH sta-
bilized) was cooled to ꢁ80 °C. Ozone gas was generated and bub-
bled through the reaction mixture (reaction gas outlet was
passed over silica gel blue for detection of ozone generation). After
the reaction mixture had turned blue, ozone flow was continued
for a further 15 min. Ozone generation was stopped and oxygen
was bubbled through the reaction mixture for ꢀ15 min or until
blue colouration had completely disappeared. Trimethylphosphine
(5 mL, 1 M in toluene) was added and the mixture was stirred for
3 h at rt. The mixture was concentrated and the resulting residue
was purified by silica gel column purification (1:5 EtOAc/PE?1:3
EtOAc/PE) to produce 13 (323 mg, 0.85 mmol) as a colourless oil
in 80% yield. Rf = 0.25 (1:3; EtOAc/PE). 1H NMR (500 MHz, CDCl3)
d 9.76 (t, J = 1.7, 2H, 2 ꢂ CH(O)-1 pentanal), 3.51 (dd, J = 4.2, 9.4,
2H, 2 ꢂ OCHH-1,3 propyl), 3.45–3.34 (m, 6H, 2 ꢂ OCHH-1,3 propyl,
2 ꢂ CH2-5 pentanal), 2.46 (dt, J = 1.7, 7.3, 4H, 2 ꢂ CH2-2 pentanal),
1.93 (s, 3H, 3 ꢂ CH Ada), 1.76–1.55 (m, 20H, 6 ꢂ CH2 Ada, 4 ꢂ CH2
pentanal), 1.31–1.25 (m, 1H, CH-2 propyl). 13C NMR (125 MHz,
CDCl3) d 202.4 (CH(O)-1 pentanal), 70.3 (CH2-5 pentanal), 68.1
(OCH2-1,3 propyl), 49.3 (CH-2 propyl), 43.6 (CH2-2 pentanal),
40.6 (CH2 Ada), 37.2 (CH2 Ada), 33.9 (Cq Ada), 29.1 (CH2-3 pen-
nyl), 29.0 (CH Ada), 25.8 (CH2-3 hexenyl). IR
mmax(thin film)/
cmꢁ1: 2902, 2849, 1640, 1450, 1366, 1110, 992, 908. HRMS: found
375.3259 [M+H]+, calculated for [C25H42O2 + H]+ 375.3258.
4.1.6. 1,10-[Adamantan-1,3-diylbis(methylene)]bis(oxy)dihex-
5-ene 12
Synthesis of 1,3-adamantane-dimethanol 10: A dry and cooled
(0 °C) solution of 1,3-adamantane-diacetic acid (9: 1 g, 4.46 mmol)
in THF (45 mL) was charged with LiAlH4 (678 mg, 17.8 mmol), stir-
red for 20 h and warmed to rt. The reaction was quenched (first
with EtOAc and then with water) and aq 1 M HCl (100 mL) was
added. The mixture was extracted with EtOAc (3 ꢂ 100 mL) and
the resulting combined organic phase was washed with sat aq
NaHCO3 (100 mL). The organic phase was dried (Na2SO4) and con-
centrated to produce 1,3-adamantane-dimethanol (10: 863 mg,
ꢀ4.4 mmol) as an off-white solid, which was used crude in the next
reaction. Rf 1,3-adamantane-dimethanol = 0.52; Rf 1,3-adaman-
tane-diacetic acid = 0.80 (19:1; EtOAc/AcOH). 1H NMR (400 MHz,
CDCl3/MeOD) d 2.96 (s, 4H, 2 ꢂ OCH2-Ada), 1.87 (s, 2H, CH-5,7
Ada), 1.43 (s, 2H, CH2-6 Ada), 1.35–1.15 (m, 8H, CH2-4,8,9,10
Ada), 1.04 (s, 2H, CH2-2 Ada). 13C NMR (100 MHz, CDCl3/MeOD) d
72.6 (HOCH2-Ada), 40.2 (CH2-2 Ada), 38.5 (CH2-4,8,9,10 Ada),
36.5 (CH2-6 Ada), 34.7 (Cq-1,3 Ada), 28.1 (CH-5,7 Ada). Synthesis
of 12: A dry cooled (0 °C) solution of 10 (196 mg, 1 mmol) and TBAI
(50 mg, 0.14 mmol) in DMF (5 mL) was charged with NaH (120 mg,
3 mmol; 60% in mineral oil). The reaction mixture was stirred for
1 h at after which 6-bromo1-hexene (0.4 mL, 3 mmol) was added.
The reaction mixture was stirred for 2 h, before warming to rt. The
reaction mixture was cooled and additional NaH (120 mg) was
tanal), 28.7 (CH Ada), 19.1 (CH2-3 pentanal). IR mmax(thin film)/
cmꢁ1
: 2901, 2848, 1723, 1451, 1367, 1109. HRMS: found
379.2843 [M+H]+, calculated for [C23H38O4 + H]+ 379.2843.
4.1.8. 3,30-{4,40-[2-(Adamantan-1-yl)propane-1,3-diyl]bis(oxy)-
bis(butane-4,1-diyl)}bis(1,2,4-trioxolane) 14 or 3,8-bis{4,40-[2-
(Adamantane-1-yl)propane-1,3-diyl]bis(oxy)bis(butane-4,1-
diyl)}-1,2,4,6,7,9-hexaoxecane 15
See note 22: Treatment of the ozonolysis reaction mixture with
DMS (0.5 mL, 6.8 mmol) for 1 h at rt resulted in a complex mixture
of products from which 14 or 15 could be isolated in ꢀ30% yield
after concentration and silica gel column purification (1:5 EtOAc/
PE?1:3 EtOAc/PE). Pure 14/15 in DCM proved stable to DMS treat-
ment (0.5 mL; 15 min). Combination of 14/15 and other minor
products and treatment with PMe3 provided 13 in good yields
(70–80%). Rf = 0.55 (1:5; EtOAc/PE). 1H NMR (400 MHz, CDCl3) d
5.18 (s, 2H, CH2-5/50 trioxolane), 5.13 (t, J = 4.9, 2H, 2 ꢂ CH-3 tri-
oxolane), 5.03 (s, 2H, CH2-5/50 trioxolane), 3.51 (dd, J = 4.2, 9.4,
2H, 2 ꢂ CHH-1,3 propyl), 3.44–3.33 (m, 6H, 2 ꢂ CHH-1,3 propyl,

T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
843
2 ꢂ CH2-4 butane), 1.93 (s, 3H, 3 ꢂ CH Ada), 1.79–1.72 (m, 4H,
2 ꢂ CH2-1 butane), 1.72–1.63 (m, 6H, 3 ꢂ CH2 Ada), 1.63–1.46
(m, 16H, 3 ꢂ CH2 Ada, 2 ꢂ CH2-3 butane), 1.55–1.49 (m, 4H,
2 ꢂ CH2-2 butane), 1.31–1.25 (m, 1H, CH-2 propyl). 13C NMR
(100 MHz, CDCl3) d 103.9 (CH-3 trioxolane), 94.1 (CH2-5/50 trioxo-
lane), 70.5 (CH2-4 butane), 68.3 (CH2-1,3 propyl), 49.5 (CH-2 pro-
pyl), 40.7 (CH2 Ada), 37.4 (CH2 Ada), 34.1 (Cq Ada), 31.0 (CH2-1
butane), 29.6 (CH2-3 butane), 28.9 (CH Ada), 20.9 (CH2-2 butane).
½
a 2D0
697.4230
ꢃ
¼ ꢁ2:4 (c 3.7, CHCl3). HRMS: found 1393.8394 [M+H]+;
[M+2H]2+
,
calculated
for
[C91H112N2O10 + H]+
1393.8390; [C91H112N2O10 + 2H]2+ 697.4231.
4.1.11. N,N0-{5,50-[2-(Adamantan-1-yl)propane-1,3-diyl]bis
(oxy)bis(pentane-5,1-diyl)}-bis(2,3,4,6-tetra-O-benzyl-L-ido-1-
deoxynojirimycin) 18
Dialdehyde 13 (159 mg, 0.42 mmol) was subjected to General
procedure A with 2,3,4,6-tetra-O-benzyl- -ido-1-deoxynojirimycin
L
4.1.9. 5,50-[Adamantan-1,3-diylbis(methylene)]bis(oxy)-
dipentanal 16
to provide 18 (322 mg, 0.23 mmol) in 55% yield as a colourless
oil after silica gel column purification. Rf = 0.70 (2:3; EtOAc/PE).
1H NMR (400 MHz, CDCl3) d 7.35–7.23 (m, 40H, CHAr Bn), 4.85 (d,
J = 11.0, 2H, 2 ꢂ CHH Bn), 4.79 (d, J = 11.0, 2H, 2 ꢂ CHH Bn), 4.70
(d, J = 11.4, 2H, 2 ꢂ CHH Bn), 4.67–4.59 (m, 6H, 2 ꢂ CHH Bn, CH2
Bn), 4.51 (d, J = 12.2, 2H, 2 ꢂ CHH Bn), 4.47 (d, J = 12.2, 2H, 2 ꢂ CHH
Bn), 3.80 (dd, J = 6.4, 10.1, 2H, 2 ꢂ H-6a), 3.72–3.63 (m, 4H, 2 ꢂ H-
4, 2 ꢂ H-6b), 3.57–3.38 (m, 8H, 2 ꢂ H-2, 2 ꢂ H-3, 2 ꢂ CH2-1,3 pro-
pyl), 3.38–3.30 (m, 6H, 2 ꢂ H-5, 2 ꢂ CH2-5 pentyl), 2.86 (dd, J = 5.2,
11.8, 2H, 2 ꢂ H-1a), 2.75–2.64 (m, 2H, 2 ꢂ NCHH-1 pentyl), 2.58–
2.46 (m, 4H, 2 ꢂ H-1b, 2 ꢂ NCHH-1 pentyl), 1.93 (s, 3H, 3 ꢂ CH
Ada), 1.71–1.58 (m, 12H, 6ꢂCH2 Ada), 1.58–1.49 (m, 4H, 2 ꢂ CH2-
4 pentyl), 1.49–1.35 (m, 4H, 2 ꢂ CH2-2 pentyl), 1.35–1.26 (m, 5H,
CH-2 propyl, 2 ꢂ CH2-3 pentyl). 13C NMR (100 MHz, CDCl3) d
139.3, 138.9, 138.8, 138.7 (4ꢂCq Bn), 128.5, 128.5, 128.4, 128.1,
127.9, 127.7, 127.7, 127.6, 127.5 (CHAr Bn), 83.3 (C-3), 80.5 (C-4),
79.1 (C-2), 75.5, 73.4, 73.2, 72.8 (4ꢂCH2 Bn), 71.1 (CH2-5,50 pentyl),
68.4 (CH2-1,3 propyl), 64.6 (C-6), 59.9 (C-5), 54.9 (NCH2-1 pentyl),
50.0 (C-1), 49.5 (CH-2 propyl), 40.8 (CH2 Ada), 37.4 (CH2 Ada), 34.2
(Cq Ada), 29.9 (CH2-4 pentyl), 29.0 (CH Ada), 28.0 (CH2-2 pentyl),
A solution of 12 (658 mg, 1.8 mmol) in DCM (75 mL; EtOH sta-
bilized) was cooled to ꢁ80 °C. Ozone gas was generated and bub-
bled through the reaction mixture (reaction gas outlet was
passed over silica gel blue for the detection of ozone generation).
After the reaction mixture had turned blue, the ozone flow was
continued for a further 15 min. Ozone generation was then stopped
and oxygen was bubbled through the reaction mixture for ꢀ15 min
or until the blue colouration had completely disappeared. Trimeth-
ylphosphine (7.5 mL, 1 M in toluene) was added and the mixture
was stirred for 20 h at 5 °C. The mixture was concentrated and
the resulting residue was purified by silica gel column purification
(1:5 EtOAc/PE?1:3 EtOAc/PE) to produce 16 (479 mg, 1.30 mmol)
as a colourless oil in 72% yield. Rf = 0.30 (1:3; EtOAc/PE). 1H NMR
(400 MHz, CDCl3) d 9.77 (t, J = 1.7, 2H, 2 ꢂ CH(O)-1 pentanal),
3.38 (t, J = 6.2, 4H, 2 ꢂ CH2-5 pentanal), 2.98 (s, 4H, 2 ꢂ OCH2-
Ada), 2.47 (dt, J = 1.7, 7.2, 4H, 2 ꢂ CH2-2 pentanal), 2.07–1.99 (m,
2H, CH-5,7 Ada), 1.74 –1.67 (m, 4H, 2 ꢂ CH2-3 pentanal), 1.63–
1.55 (m, 6H, CH2-6 Ada, 2 ꢂ CH2-4 pentanal), 1.53–1.38 (m, 8H,
CH2-4,8,9,10 Ada), 1.30 (s, 2H, CH2-2 Ada). 13C NMR (100 MHz,
CDCl3) d 202.7 (CH(O)-1 pentanal), 81.6 (OCH2-Ada), 70.9 (CH2-5
pentanal), 43.6 (CH2-2 pentanal), 41.8 (CH2-2 Ada), 39.4 (CH2-
4,8,9,10 Ada), 36.8 (CH2-6 Ada), 34.5 (Cq-1,3 Ada), 29.0 (CH2-4 pen-
24.2 (CH2-3 pentyl). IR
m
max(thin film)/cmꢁ1: 3031, 2902, 2851,
1496, 1454, 1364, 1093, 1027, 734, 697. ½a D20
¼ ꢁ21:8 (c 1.8,
ꢃ
CHCl3). HRMS: found 1393.8392 [M+H]+; 697.4230 [M+2H]2+, cal-
culated for [C91H112N2O10 + H]+ 1393.8390; [C91H112N2O10 + 2H]2+
697.4231.
tanal), 28.3 (CH-5,7 Ada), 19.0 (CH2-3 pentanal). IR mmax(thin film)/
cmꢁ1: 2898, 2847, 1722, 1454, 1366, 1126, 1103. HRMS: found
365.2693 [M+H]+, calculated for [C22H36O4 + H]+ 365.2686.
4.1.12. N,N0-{5,50-[Adamantan-1,3-diylbis(methylene)]bis(oxy)-
bis(pentane-5,1-diyl)}-bis(2,3,4,6-tetra-O-benzyl-1-
deoxynojirimycin) 19
4.1.10. N,N0-{5,50-[2-(Adamantan-1-yl)propane-1,3-diyl]bis-
(oxy)bis(pentane-5,1-diyl)}-bis(2,3,4,6-tetra-O-benzyl-1-
deoxynojirimycin) 17
Dialdehyde 16 (119 mg, 0.33 mmol) was subjected to general
procedure A with 2,3,4,6-tetra-O-benzyl-1-deoxynojirimycin to
provide 19 (282 mg, 0.20 mmol) in 62% yield as a colourless oil
after silica gel column purification. Rf = 0.57 (2:3; EtOAc/PE). 1H
NMR (400 MHz, CDCl3) d 7.36–7.20 (m, 36H, CHAr Bn), 7.15–7.10
(m, 4H, CHAr Bn), 4.95 (d, J = 11.1, 2H, 2 ꢂ CHH Bn), 4.87 (d,
J = 10.8, 2H, 2 ꢂ CHH Bn), 4.81 (d, J = 11.1, 2H, 2 ꢂ CHH Bn), 4.69
(d, J = 11.6, 2H, 2 ꢂ CHH Bn), 4.64 (d, J = 11.6, 2H, 2 ꢂ CHH Bn),
4.49 (d, J = 12.3, 2H, 2 ꢂ CHH Bn), 4.45 (d, J = 12.3, 2H, 2 ꢂ CHH
Bn), 4.41 (d, J = 10.8, 2H, 2 ꢂ CHH Bn), 3.70–3.63 (m, 4H, 2 ꢂ H-2,
2 ꢂ H-6a), 3.60 (dd, J = 9.3, 2H, 2 ꢂ H-4), 3.53 (dd, J = 2.1, 10.3,
2H, 2 ꢂ H-6b), 3.45 (dd, J = 9.1, 2H, 2 ꢂ H-3), 3.33 (t, J = 6.5, 4H,
2 ꢂ CH2-5 pentyl), 3.09 (dd, J = 4.8, 11.1, 2H, 2 ꢂ H-1a), 2.99 (s,
4H, 2 ꢂ OCH2-Ada), 2.72–2.63 (m, 2H, 2 ꢂ NCHH-1 pentyl), 2.62–
2.52 (m, 2H, 2 ꢂ NCHH-1 pentyl), 2.29 (dt, J = 2.1, 9.6, 2H, 2 ꢂ H-
5), 2.23 (t, J = 10.8, 2H, 2 ꢂ H-1b), 2.05 (s, 2H, CH-5,7 Ada), 1.62
(s, 2H, CH2-6 Ada), 1.56–1.43 (m, 12H, CH2-4,8,9,10 Ada, 2 ꢂ CH2-
4 pentyl), 1.43–1.33 (m, 4H, 2 ꢂ CH2-2 pentyl), 1.31 (s, 2H, CH2-2
Ada), 1.28–1.12 (m, 4H, 2 ꢂ CH2-3 pentyl). 13C NMR (100 MHz,
CDCl3) d 139.2, 138.7, 138.7, 137.9 (4ꢂCq Bn), 128.6, 128.5, 128.4,
128.4, 128.0, 127.9, 127.7, 127.6, 127.5 (CHAr Bn), 87.5 (C-3), 81.8
(OCH2-Ada), 78.8, 78.7 (C-2, C-4), 75.4, 75.3, 73.6, 72.9 (4 ꢂ CH2
Bn), 71.7 (CH2-5 pentyl), 65.4 (C-6), 63.8 (C-5), 54.6 (C-1), 52.5
(NCH2-1 pentyl), 42.0 (CH2-2 Ada), 39.6 (CH2-4,8,9,10 Ada), 36.9
(CH2-6 Ada), 34.7 (Cq-1,3 Ada), 29.6 (CH2-4 pentyl), 28.5 (CH-5,7
Dialdehyde 13 (159 mg, 0.42 mmol) was subjected to General
procedure A with 2,3,4,6-tetra-O-benzyl-1-deoxynojirimycin to
provide 17 (343 mg, 0.25 mmol) in 60% yield as a colourless oil
after silica gel column purification. Rf = 0.58 (2:3; EtOAc/PE). 1H
NMR (400 MHz, CDCl3) d 7.36–7.24 (m, 36H, CHAr Bn), 7.15–7.09
(m, 4H, CHAr Bn), 4.95 (d, J = 11.1, 2H, 2 ꢂ CHH Bn), 4.87 (d,
J = 10.9, 2H, 2 ꢂ CHH Bn), 4.81 (d, J = 11.1, 2H, 2 ꢂ CHH Bn), 4.68
(d, J = 11.6, 2H, 2 ꢂ CHH Bn), 4.64 (d, J = 11.6, 2H, 2 ꢂ CHH Bn),
4.46 (s, 4H, 2 ꢂ CH2 Bn), 4.40 (d, J = 10.9, 2H, 2 ꢂ CHH Bn), 3.70–
3.62 (m, 4H, 2 ꢂ H-2, 2 ꢂ H-6a), 3.59 (dd, J = 9.3, 9.5, 2H, 2 ꢂ H-
4), 3.56–3.38 (m, 8H, 2 ꢂ H-3, 2 ꢂ H-6b, 2 ꢂ CH2-1,3 propyl),
3.38–3.27 (m, 4H, 2 ꢂ CH2-5 pentyl), 3.08 (dd, J = 4.8, 11.1, 2H,
2 ꢂ H-1a), 2.72–2.63 (m, 2H, 2 ꢂ NCHH-1 pentyl), 2.63–2.54 (m,
2H, 2 ꢂ NCHH-1 pentyl), 2.30 (dt, J = 2.1, 9.5, 2H, 2 ꢂ H-5), 2.23
(dd, J = 10.5, 11.1, 2H, 2 ꢂ H-1b), 1.94 (s, 3H, 3 ꢂ CH Ada), 1.73–
1.59 (m, 12H, 6 ꢂ CH2 Ada), 1.56–1.46 (m, 4H, 2 ꢂ CH2-4 pentyl),
1.45–1.17 (m, 9H, CH-2 propyl, 2 ꢂ CH2-2 pentyl, 2 ꢂ CH2-3 pen-
tyl). 13C NMR (100 MHz, CDCl3) d 139.2, 138.8, 138.8, 138.0
(4 ꢂ Cq Bn), 128.6, 128.6, 128.5, 128.0, 128.0, 127.8, 127.7, 127.6
(CHAr Bn), 87.6 (C-3), 78.8, 78.7 (C2, C-4), 75.5, 75.3, 73.6, 72.9
(4ꢂCH2 Bn), 71.1, 71.0 (CH2-5,50 pentyl), 68.5 (CH2-1,3 propyl),
65.5 (C-6), 63.9 (C-5), 54.7 (C-1), 52.6 (NCH2-1,10 pentyl), 49.5
(CH-2 propyl), 40.8 (CH2 Ada), 37.5 (CH2 Ada), 34.2 (Cq Ada), 29.9
(CH2-4,40 pentyl), 29.0 (CH Ada), 24.5, 24.4 (CH2-3,30 pentyl),
Ada), 24.2 (CH2-3 pentyl), 23.5 (CH2-2 pentyl). IR mmax(thin film)/
23.6, 23.5 (CH2-2,20 pentyl). IR
2853, 1495, 1454, 1361, 1274, 1208, 1093, 1027, 734, 697.
m
max(thin film)/cmꢁ1: 3031, 2903,
cmꢁ1: 3030, 2900, 2849, 1495, 1454, 1360, 1208, 1094, 1027,
734, 696. ½a 2D0
¼ ꢁ3:0 (c 2.4, CHCl3). HRMS: found 1379.8234
ꢃ

844
T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
[M+H]+; 690.4150 [M+2H]2+, calculated for [C90H110N2O10 + H]+
1379.8233; [C90H110N2O10 + 2H]2+ 690.4153.
2H, 2 ꢂ H-3), 3.55 (dd, J = 4.2, 9.5, 2H, 2 ꢂ CHH-1,3 propyl), 3.50
(s, 2H, 2 ꢂ H-5), 3.48–3.36 (m, 8H, 2 ꢂ CHH-1,3 propyl, 2 ꢂ CH2-5
pentyl), 3.34–3.24 (m, 4H, 2 ꢂ CH2-1, 2 ꢂ CH2-1 pentyl), 1.94 (s,
3H, 3 ꢂ CH Ada), 1.90–1.76 (m, 4H, 2 ꢂ CH2-2 pentyl), 1.76–1.59
(m, 16H, 6 ꢂ CH2 Ada, 2 ꢂ CH2-4 pentyl), 1.51–1.41 (m, 4H,
2 ꢂ CH2-3 pentyl), 1.31–1.25 (m, 1H, CH-2 propyl). 13C NMR
(100 MHz, MeOD) d 72.0 (C-4), 71.7 (CH2-5 pentyl), 69.4 (CH2-1,3
propyl), 69.2 (C-2, C-3), 63.7 (C-5), 60.2 (C-6 collapsed), 55.2
(CH2-1 pentyl), 53.9 (C-1), 50.8 (CH-2 propyl), 41.8 (CH2 Ada),
38.4 (CH2 Ada), 35.2 (Cq Ada), 30.4 (CH2-4 pentyl), 30.3 (CH Ada),
4.1.13. N,N0-{5,50-[Adamantan-1,3-diylbis(methylene)]bis(oxy)-
bis(pentane-5,1-diyl)}-bis(2,3,4,6-tetra-O-benzyl-L-ido-1-
deoxynojirimycin) 20
Dialdehyde 16 (119 mg, 0.33 mmol) was subjected to general
procedure A with 2,3,4,6-tetra-O-benzyl- -ido-1-deoxynojirimycin
L
to provide 20 (293 mg, 0.21 mmol) in 64% yield as a colourless
oil after silica gel column purification. Rf = 0.69 (2:3; EtOAc/PE).
1H NMR (400 MHz, CDCl3) d 7.35–7.23 (m, 40H, CHAr Bn), 4.85 (d,
J = 11.0, 2H, 2 ꢂ CHH Bn), 4.79 (d, J = 11.0, 2H, 2 ꢂ CHH Bn), 4.70
(d, J = 11.5, 2H, 2 ꢂ CHH Bn), 4.67–4.60 (m, 6H, 2 ꢂ CHH Bn, CH2
Bn), 4.52 (d, J = 12.2, 2H, 2 ꢂ CHH Bn), 4.48 (d, J = 12.2, 2H, 2 ꢂ CHH
Bn), 3.80 (dd, J = 6.4, 10.1, 2H, 2 ꢂ H-6a), 3.70 (dd, J = 2.4, 10.1, 2H,
2 ꢂ H-6b), 3.66 (dd, J = 5.9, 9.4, 2H, 2 ꢂ H-4), 3.58–3.52 (m, 2H,
2 ꢂ H-2), 3.52–3.46 (m, 2H, 2 ꢂ H-3), 3.38–3.31 (m, 5H,, 2 ꢂ H-5,
2 ꢂ CH2-5 pentyl), 2.98 (s, 4H, 2 ꢂ OCH2-Ada), 2.86 (dd, J = 5.3,
11.9, 2H, 2 ꢂ H-1a), 2.70 (ddd, J = 6.5, 8.7, 12.5, 2H, 2 ꢂ NCHH-1
pentyl), 2.56–2.48 (m, 4H, 2 ꢂ H-1a, 2 ꢂ NCHH-1 pentyl), 2.03 (s,
2H, CH-5,7 Ada), 1.68–1.35 (m, 18H, CH2-4,6,8,9,10 Ada, 2 ꢂ CH2-
24.8 (CH2-3 pentyl), 24.3 (CH2-2 pentyl). IR m :
max(thin film)/cmꢁ1
3314, 2902, 2849, 1447, 1065. ½a D20
¼ þ11:4 (c 2.6, MeOH). HRMS:
ꢃ
found 673.4633 [M+H]+, calculated for [C35H64N2O10 + H]+
673.4634.
4.1.16. N,N0-{5,50-[Adamantan-1,3-diylbis(methylene)]bis(oxy)-
bis(pentane-5,1-diyl)}-bis(1-deoxynojirimycin) 23
Compound 19 (624 mg, 0.45 mmol) was subjected to general
procedure B to provide 23 (177 mg, 0.27 mmol) in 60% yield as a
colourless oil after silica gel column purification. Rf = 0.40 (19:1;
EtOAc/NH4OH).1H NMR (400 MHz, MeOD) d 4.04 (dd, J = 1.7, 12.5,
2H, 2 ꢂ H-6a), 3.92 (dd, J = 2.7, 12.5, 2H, 2 ꢂ H-6a), 3.72 (ddd,
J = 4.9, 9.3, 10.9, 2H, 2 ꢂ H-2), 3.57 (dd, J = 9.1, 9.6, 2H, 2 ꢂ H-4),
3.47–3.33 (m, 8H, 2 ꢂ CH2-5 pentyl, 2 ꢂ H-1a, 2 ꢂ H-3), 3.27–
3.15 (m, 2H, 2 ꢂ NCHH-1 pentyl), 3.14–3.03 (m, 2H, 2 ꢂ NCHH-1
pentyl), 3.02 (s, 4H, 2 ꢂ OCH2-Ada), 2.86 (d, J = 9.8, 2H, 2 ꢂ H-5),
2.81 (dd, J = 10.5, 11.5, 2H, 2 ꢂ H-1b), 2.02 (s, 2H, 2 ꢂ CH-5,7
Ada), 1.72 (s, 4H, 2 ꢂ CH2-2 pentyl), 1.67–1.58 (m, 6H, CH2-6
Ada, 2 ꢂ CH2-4 pentyl), 1.56–1.37 (m, 12H, CH2-4,8,9,10 Ada,
2 ꢂ CH2-3 pentyl), 1.32 (s, 2H, CH2-2 Ada). 13C NMR (100 MHz,
MeOD) d 82.8 (OCH2-Ada), 78.6 (C-3), 72.3 (CH2-5 pentyl), 69.8
(C-4), 68.7 (C-2), 67.4 (C-5), 56.6 (C-6), 55.6 (C-1), 54.1 (NCH2-1
pentyl), 43.0 (CH2-2 Ada), 40.6 (CH2-4,8,9,10 Ada), 38.0 (CH2-6
Ada), 35.7 (Cq-1,3 Ada), 30.3 (CH2-4 pentyl), 29.8 (CH-5,7 Ada),
2
pentyl, 2 ꢂ CH2-4 pentyl), 1.35–1.22 (m, 6H, CH-2 Ada,
2 ꢂ CH2-3 pentyl). 13C NMR (100 MHz, CDCl3) d 139.3, 138.8,
138.8, 138.7 (4ꢂCq Bn), 128.5, 128.5, 128.4, 128.4, 128.1, 127.9,
127.7, 127.7, 127.6, 127.5 (CHAr Bn), 83.3 (C-3), 81.8 (OCH2-Ada),
80.5 (C-4), 79.1 (C-2), 75.5, 73.4, 73.2, 72.8 (4ꢂCH2 Bn), 71.8
(CH2-5 pentyl), 64.5 (C-6), 59.9 (C-5), 54.8 (NCH2-1 pentyl), 50.0
(C-1), 42.1 (CH2-2 Ada), 39.6 (CH2-4,8,9,10 Ada), 36.9 (CH2-6
Ada), 34.7 (Cq-1,3 Ada), 29.6 (CH2-4 pentyl), 28.5 (CH-5,7 Ada),
28.0 (CH2-2 pentyl), 24.0 (CH2-3 pentyl). IR m :
max(thin film)/cmꢁ1
3030, 2900, 2849, 1496, 1454, 1363, 1207, 1094, 1027, 734, 697.
½
a 2D0
690.4151
ꢃ
¼ ꢁ25:9 (c 2.1, CHCl3). HRMS: found 1379.8236 [M+H]+;
[M+2H]2+
,
calculated
for
[C90H110N2O10 + H]+
1379.8233; [C90H110N2O10 + 2H]2+ 690.4153.
24.8 (CH2-3 pentyl), 24.3 (CH2-2 pentyl). IR m :
max(thin film)/cmꢁ1
4.1.14. N,N0-{5,50-[2-(Adamantan-1-yl)propane-1,3-diyl]bis-
(oxy)bis(pentane-5,1-diyl)}-bis(1-deoxynojirimycin) 21
3296, 2899, 2848, 1453, 1369, 1102, 1029. ½a D20
¼ ꢁ3:6 (c 3.6,
ꢃ
MeOH). HRMS: found 659.4475 [M+H]+, calculated for
[C34H62N2O10 + H]+ 659.4477.
Compound 17 (377 mg, 0.27 mmol) was subjected to general
procedure B to provide 21 (144 mg, 0.21 mmol) in 79% yield as a
colourless oil after silica gel column purification. Rf = 0.40 (19:1;
EtOAc/NH4OH); Rf = 0.05 (1:1; EtOAc/MeOH+5%NH4OH). 1H NMR
(400 MHz, MeOD) d 3.93 (dd, J = 2.4, 12.1, 2H, 2 ꢂ H-6a), 3.88
(dd, J = 2.7, 12.1, 2H, 2 ꢂ H-6b), 3.59–3.53 (m, 4H, 2 ꢂ H-2,
2 ꢂ CHH-1,3 propyl), 3.49–3.37 (m, 8H, 2 ꢂ H-4, 2 ꢂ CHH-1,3 pro-
pyl, 2 ꢂ CH2-5 pentyl), 3.22 (dd, J = 9.1, 9.1, 2H, 2 ꢂ H-3), 3.14 (dd,
J = 4.8, 11.5, 2H, 2 ꢂ H-1a), 3.03–2.93 (m, 2H, 2 ꢂ NCHH-1 pentyl),
2.84–2.74 (m, 2H, 2 ꢂ NCHH-1 pentyl), 2.49–2.40 (m, 4H, 2 ꢂ H-1b,
2 ꢂ H-5), 1.94 (s, 3H, 3 ꢂ CH Ada), 1.78–1.64 (m, 12H, 6ꢂCH2 Ada),
1.64–1.56 (m, 8H, 2 ꢂ CH2-2 pentyl, 2 ꢂ CH2-3 pentyl), 1.45–1.35
(m, 4H, 2 ꢂ CH2-3 pentyl), 1.31–1.25 (m, 1H, CH-2 propyl). 13C
NMR (100 MHz, MeOD) d 79.9 (C-3), 71.9 (CH2-5 pentyl), 71.1 (C-
4), 69.9 (C-2), 69.4 (CH2-1,3 propyl), 67.5 (C-5), 58.2 (C-6), 56.9
(C-1), 54.0 (NCH2-1 pentyl), 50.8 (CH-2 propyl), 41.9 (CH2 Ada),
38.4 (CH2 Ada), 35.2 (Cq Ada), 30.7 (CH2-4 pentyl), 30.3 (CH Ada),
4.1.17. N,N0-{5,50-[Adamantan-1,3-diylbis(methylene)]bis(oxy)-
bis(pentane-5,1-diyl)}-bis(L-ido-1-deoxynojirimycin) 24
Compound 20 (611 mg, 0.44 mmol) was subjected to general
procedure B to provide 24 (159 mg, 0.24 mmol) in 55% yield as a
colourless oil after silica gel column purification. Rf = 0.45 (19:1;
EtOAc:NH4OH). 1H NMR (400 MHz, MeOD) d 4.08–3.99 (m, 6H,
2 ꢂ CH2-6, 2 ꢂ H-4), 3.99–3.93 (m, 2H, 2 ꢂ H-2), 3.91–3.81 (m,
2H, 2 ꢂ H-3), 3.59–3.50 (m, 2H, 2 ꢂ H-5), 3.49–3.32 (m, 12H,
2 ꢂ NCH2-1 pentyl, 2 ꢂ CH2-5 pentyl, 2 ꢂ CH2-1), 3.01 (s, 4H,
2 ꢂ OCH2-Ada), 2.02 (s, 2H, 2 ꢂ CH-5,7 Ada), 1.92–1.71 (m, 4H,
2 ꢂ CH2-2 pentyl), 1.69–1.58 (m, 6H, CH2-6 Ada, 2 ꢂ CH2-4 pentyl),
1.57–1.41 (m, 12H, CH2-4,8,9,10 Ada, 2 ꢂ CH2-3 pentyl), 1.32 (s,
2H, CH2-2 Ada). 13C NMR (100 MHz, MeOD) d 82.9 (OCH2-Ada),
72.3 (CH2-5 pentyl), 71.7 (C-4), 68.9 (C-2, C-3), 63.7 (C-5), 60.5
(C-6 collapsed), 55.2 (CH2-1 pentyl), 54.0 (C-1), 43.0 (CH2-2 Ada),
40.6 (CH2-4,8,9,10 Ada), 38.0 (CH2-6 Ada), 35.7 (Cq-1,3 Ada), 30.2
(CH2-4 pentyl), 29.8 (CH-5,7 Ada), 24.7 (CH2-3 pentyl), 24.1 (CH2-
25.3 (CH2-3 pentyl), 24.8 (CH2-2 pentyl). IR m :
max(thin film)/cmꢁ1
3344, 2902, 2849, 1448, 1370, 1108, 1032. ½a D20
¼ ꢁ8:8 (c 1.9,
ꢃ
MeOH). HRMS: found 673.4633 [M+H]+, calculated for
[C35H64N2O10 + H]+ 673.4634.
2 pentyl). IR m
max(thin film)/cmꢁ1: 3312, 2899, 2848, 1450, 1101,
1027.
½
a 2D0
ꢃ
¼ þ13:2 (c 3.1, MeOH). HRMS: found 659.4476
[M+H]+, calculated for [C34H62N2O10 + H]+ 659.4477.
4.1.15. N,N0-{5,50-[2-(Adamantan-1-yl)propane-1,3-diyl]bis-
(oxy)bis(pentane-5,1-diyl)}-bis(
L
-ido-1-deoxynojirimycin) 22
4.1.18. 5-(Adamantan-1-yl-ethoxy)-1-bromo-pentane 26
A solution of PPh3 (5.53 g, 21.1 mol) in CH3CN (50 mL) was
added to a solution of 5-(adamantan-1-yl-ethoxy)-1-pentanol
(25: 1.87 g, 7.0 mmol) and carbontetrabromide (4.66 g, 14.1 mmol)
in CH3CN (150 mL). The reaction mixture was refluxed for 3 h and
subsequently concentrated. The residue was purified by silica gel
Compound 18 (435 mg, 0.31 mmol) was subjected to general
procedure B to provide 22 (153 mg, 0.23 mmol) in 73% yield as a
colourless oil after silica gel column purification. Rf = 0.45 (19:1;
EtOAc/NH4OH). 1H NMR (400 MHz, MeOD) d 4.05–3.95 (m, 6H,
2 ꢂ CH2-6, 2 ꢂ H-4), 3.95–3.90 (m, 2H, 2 ꢂ H-2), 3.84–3.79 (m,

T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
845
column purification (100% PE?1:19 EtOAc/PE) to provide 26 (2.01
References
g, 6.13 mmol) in 87% yield as a colourless oil. 5-(Adamantan-1-yl-
ethoxy)-1-pentanol was obtained via the same route as previously
described for 5-(adamantan-1-yl-methoxy)-1-pentanol,6 but now
with commercially available 2-adamantaneethanol. Rf = 0.15 (1:9;
toluene/PE). 1H NMR (400 MHz, CDCl3) d 3.42 (dt, J = 6.9, 13.2,
6H, CH2-1 pentyl, CH2-5 pentyl, OCH2 ethoxy), 1.93 (s, 3H,
3 ꢂ CH Ada), 1.92–1.84 (m, 2H, CH2-2 pentyl), 1.73–1.61 (m, 6H,
3 ꢂ CH2 Ada), 1.61–1.54 (m, 2H, CH2-4 pentyl), 1.54–1.45 (m, 8H,
3 ꢂ CH2 Ada, CH2-3 pentyl), 1.37 (t, J = 7.5, 2H, CH2-Ada ethoxy).
13C NMR (100 MHz, CDCl3) d 70.6 (CH2-5 pentyl), 66.9 (OCH2 eth-
oxy), 43.8 (CH2-Ada ethoxy), 42.9 (CH2 Ada), 37.3 (CH2 Ada), 33.9
(CH2-1 pentyl), 32.8 (CH2-2 pentyl), 31.8 (Cq Ada), 29.1 (CH2-4 pen-
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1312.
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M. O.; Hammer, R. E.; Hamra, F. K.; Matern, S.; Russell, D. W. J. Clin. Invest. 2006,
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Strijland, A.; Aerts, J. M. F. G.; Overkleeft, H. S. J. Org. Chem. 2007, 72, 1088–
1097.
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Dev. 2008, 12, 414–423.
tyl), 28.8 (CH Ada), 25.2 (CH2-3 pentyl). IR m :
max(thin film)/cmꢁ1
2897, 2845, 1728, 1450, 1362, 1344, 1271, 1109, 968.
8. Overkleeft, H. S.; Renkema, G. H.; Neele, J.; Vianello, P.; Hung, I. O.; Strijland, A.;
van der Burg, A. M.; Koomen, G. J.; Pandit, U. K.; Aerts, J. M. F. G. J. Biol. Chem.
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9. Butters, T. D. Curr. Opin. Chem. Biol. 2007, 11, 412–418.
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Dis. 2006, 29, 449–456.
11. Butters, T. D.; Dwek, R. A.; Fleet, G.; Orchard, M. G.; Platt, F. M., WO 02/055498
A1, 2002.
12. Butters, T. D. In Iminosugars: From Synthesis to Therapeutic Applications;
Compain, P., Martin, O. R., Eds.; Iminosugar inhibitors for substrate reduction
therapy for the lysosomal glycosphingolipidoses; Wiley-VCH: New York, 2007;
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13. Wennekes, T.; Meijer, A. J.; Boot, R. G.; Groen, A. K.; Groener, J. E.; van Eijk, M.;
Ottenhoff, R.; Bijl, N.; Song, H.; van den Berg, R. J. B. H. N.; van der Marel, G. A.;
Overkleeft, H. S.; Aerts, J. M. F. G., submitted for publication.
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2755–2794.
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1971.
4.1.19. N-[5-(Adamantan-1-yl-ethoxy)-pentyl]-1-
deoxynojirimycin 27
Bromide 26 (148 mg, 0.45 mmol) was added to a mixture of 1-
deoxynojirimycin (49 mg, 0.3 mmol; synthesis as reported previ-
ously13) and K2CO3 (124 mg, 0.9 mmol) in DMF (1.5 mL). The mix-
ture was heated at 90 °C for 48 h. The mixture was filtered over a
glass microfibre filter and the filtrate was concentrated. Silica gel
column purification (100% EtOAc?3:1 EtOAc:MeOH+5%NH4OH)
of the residue provided 27 (70 mg, 0.17 mmol) in 58% yield as a
colourless oil. Rf = 0.45 (1:2; MeOH/EtOAc + NH4OH). 1H NMR
(600 MHz, MeOD) d 3.88–3.83 (m, 2H, CH2-6), 3.51–3.45 (m, 3H,
H-2, OCH2 ethoxy), 3.42 (t, J = 6.5, 2H, CH2-5 pentyl), 3.36 (dd,
J = 9.3, 1H, H-4), 3.14 (dd, J = 9.1, 1H, H-3), 3.01 (dd, J = 4.8, 11.2,
1H, H-1a), 2.86–2.79 (m, 1H, CHH-1 pentyl), 2.65–2.58 (m, 1H,
CHH-1 pentyl), 2.21 (dd, J = 10.8, 1H, H-1b), 2.15 (d, J = 7.6, 1H,
H-5), 1.93 (s, 3H, 3 ꢂ CH Ada), 1.71 (dd, J = 11.6, 41.9, 6H,
3 ꢂ CH2 Ada), 1.62–1.56 (m, 2H, CH2-4 pentyl), 1.56 (d, J = 2.6,
6H, 3 ꢂ CH2 Ada), 1.55–1.49 (m, 2H, CH2-2 pentyl), 1.39–1.31 (m,
4H, CH2-3 pentyl, CH2-Ada ethoxy). 13C NMR (150 MHz, MeOD) d
80.6 (C-3), 72.0 (C-4), 71.9 (CH2-5 pentyl), 70.7 (C-2), 67.9 (OCH2
ethoxy), 67.5 (C-5), 59.3 (C-6), 57.7 (C-1), 53.9 (CH2-1 pentyl),
44.9 (CH2-Ada ethoxy), 44.0 (CH2 Ada), 38.3(CH2 Ada), 32.9 (Cq
Ada), 30.8 (CH2-4 pentyl), 30.3 (CH Ada), 25.4 (CH2-3 pentyl),
18. Diot, J.; Garcia-Moreno, M. I.; Gouin, S. G.; Mellet, C. O.; Haupt, K.; Kovensky, J.
Org. Biomol. Chem. 2009, 7, 357–363.
19. van den Broek, L. A. G. M.; Vermaas, D. J.; van Kemenade, F. J.; Tan, M. C. C. A.;
Rotteveel, F. T. M.; Zandberg, P.; Butters, T. D.; Miedema, F.; Ploegh, H. L.; van
Boeckel, C. A. A. Recl. Trav. Chim. Pays-Bas. 1994, 113, 507–516.
20. Nasr, K.; Pannier, N.; Frangioni, J. V.; Maison, W. J. Org. Chem. 2008, 73, 1056–
1060.
21. Pfeffer, P. E.; Kinsel, E.; Silbert, L. S. J. Org. Chem. 1972, 37, 1256–1258.
22. Note on the isolated ozonide 14 or 15: Isolation and NMR analysis of the major
product after ozonolysis of 11 and DMS treatment confirm the general ozonide
structure. However, ozonide 14 is the expected ozonide obtained after
ozonolysis of a terminal alkene and should be susceptible to DMS treatment.
There is precedent by Odinokov et al.34 for the formation of a bis-substituted
1,2,4,6,7,9-hexaoxecane such as 15 after ozonolysis of terminal dialkenes and
15 might be less susceptible to DMS treatment.
25.1 (CH2-2 pentyl). IR
m
max(thin film)/cmꢁ1: 3360, 2899, 2844,
1673, 1456, 1097, 1013, 980. ½a D20
¼ ꢁ16:6 (c 0.8, MeOH). HRMS:
ꢃ
found 412.3058 [M+H]+, calculated for [C23H41NO5 + H]+ 412.3057.
4.1.20. TFA salt of N-[5-(Adamantan-1-yl-ethoxy)-pentyl]-
L-ido-
1-deoxynojirimycin 28ꢄTFA
O
Bromide 26 (148 mg, 0.45 mmol) was added to a mixture of L-
O
O
O
O
ido-1-deoxynojirimycin (49 mg, 0.3 mmol; synthesis as reported
previously13) and K2CO3 (124 mg, 0.9 mmol) in DMF (1.5 mL).
The mixture was heated at 90 °C for 48 h. The mixture was filtered
over a glass microfibre filter and the filtrate was concentrated. Sil-
ica gel column purification (100% EtOAc?3:1 EtOAc/MeOH + 5%N-
H4OH) of the residue provided 28 (53 mg, 0.13 mmol) in 43% yield
as a colourless oil. Rf = 0.35 (2:3; MeOH/EtOAc + NH4OH). 1H NMR
(400 MHz, D2O) d 4.06–3.46 (m, 4H, CH2-6, H-2, H-3, H-4), 3.46–
3.36 (m, 1H, H-5), 3.36–3.16 (m, 6H, CH2-5 pentyl, CH2-1 pentyl,
OCH2 ethoxy), 3.16–3.03 (m, 2H, CH2-1), 1.74 (s, 3H, 3 ꢂ CH Ada),
1.69–1.27 (m, 8H, 6ꢂCH2 Ada, 2 ꢂ CH2 pentyl), 1.27–1.04 (m, 3H,
CH2 pentyl, CH2-Ada ethoxy). 13C NMR (100 MHz, D2O) d 162.5,
162.1, 161.8, 161.4 (C@O TFA), 120.2, 117.3, 114.4, 111.5 (CF3
TFA), 70.2 (CH2-5 pentyl), 69.4, 67.2, 66.5 (C-2, C-3, C-4), 66.4
(OCH2 ethoxy), 61.3 (C-5), 58.5 (C-6), 53.5 (C-1), 52.2 (CH2-1 pen-
tyl), 42.9 (CH2-Ada ethoxy), 42.3 (CH2 Ada), 36.9 (CH2 Ada), 31.2
(Cq Ada), 28.6 (CH2 Ada, CH2 pentyl), 22.7, 22.1 (2 ꢂ CH2 pentyl).
15
O
O
O
14
O
O
4(CH2)
(CH2)4
O
O
O
O
O
O
23. Fleet, G. W. J.; Karpas, A.; Dwek, R. A.; Fellows, L. E.; Tyms, A. S.; Petursson,
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25. Davis, M. C.; Dahl, J. E. P.; Carlson, R. M. K. Synth. Commun. 2008, 38, 1153–
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R. M. K.; Fokin, A. A.; Schreiner, P. R. Eur. J. Org. Chem. 2007, 4738–
4745.
IR
m
max(thin film)/cmꢁ1: 3362, 2902, 2848, 1670, 1450, 1182,
27. Schreiner, P. R.; Fokina, N. A.; Tkachenko, B. A.; Hausmann, H.; Serafin, M.;
Dahl, J. E. P.; Liu, S. G.; Carlson, R. M. K.; Fokin, A. A. J. Org. Chem. 2006, 71,
6709–6720.
1139, 1067, 1024, 837. ½a D20
¼ þ11:9 (c 0.8, MeOH). HRMS: found
ꢃ
412.3055 [M+H]+, calculated for [C23H41NO5 + H]+ 412.3057.

846
T. Wennekes et al. / Tetrahedron: Asymmetry 20 (2009) 836–846
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