Carbohydrate transition state mimics: Synthesis of imidazolo-pyrrolidinoses as potential nectrisine surrogates

Article abstract of DOI:10.1016/S0968-0896(03)00402-4

The syntheses of four glyco-imidazoles, which are pentose-derivatives belonging to the D-series, as well as the syntheses of their L-enantiomers, are reported. Starting from the known linear xylo, lyxo, arabino, and ribo imidazolo-pentoses in both the L- and the D-series, intramolecular Walden inversion led to the corresponding arabino, ribo, xylo, and lyxo pyrrolidinopentoses in the D- and the L-series, respectively, protection and deprotection steps being unavoidable prerequisites. The structures and configurations of all eight pyrrolidinopentoses were determined unambiguously, by a combination of ¹H/¹³C NMR spectroscopy, circular dichroism and [α]_D values, in conjunction with single-crystal X-ray diffraction analysis of the L-xylo stereoisomer. Examination of the inhibitory properties of these imidazolo-pyrrolidinoses against six commonly encountered glycosidases led to the conclusion that by and large the L-stereoisomers are inactive, whereas three out the four D-stereoisomers proved to be poor to moderate inhibitors. It appears therefore that the most basic N(1) atom is not located in an optimal topology to be protonated easily inside the enzyme's active site.

Full text of DOI:10.1016/S0968-0896(03)00402-4

Bioorganic & Medicinal Chemistry 11 (2003) 3559–3568
Carbohydrate Transition State Mimics: Synthesis of Imidazolo-
Pyrrolidinoses as Potential Nectrisine Surrogates
Theophile Tschamber,a Francois Gessier,a Estelle Dubost,a Jeffery Newsome,a
Celine Tarnus,a Josiane Kohler,b Markus Neuburgerc and Jacques Streitha,*
a
´
´
Ecole Nationale Superieure de Chimie, Universite de Haute-Alsace, 3, rue Alfred Werner, F-68093 Mulhouse, France
bOptical Spectroscopy, F. Hoffmann-La Roche Ltd, Grenzacherstrasse, CH-4070 Basle, Switzerland
cInstitut fur anorganische Chemie, Universitat Basel, Spitalstrasse 51, CH-4056 Basle, Switzerland
¨
¨
Received 31 January 2003; accepted 5 June 2003
Abstract—The syntheses of four glyco-imidazoles, which are pentose-derivatives belonging to the d-series, as well as the syntheses
of their l-enantiomers, are reported. Starting from the known linear xylo, lyxo, arabino, and ribo imidazolo-pentoses in both the l-
and the d-series, intramolecular Walden inversion led to the corresponding arabino, ribo, xylo, and lyxo pyrrolidinopentoses in the
d- and the l-series, respectively, protection and deprotection steps being unavoidable prerequisites. The structures and configur-
1
ations of all eight pyrrolidinopentoses were determined unambiguously, by a combination of H/13C NMR spectroscopy, circular
dichroism and [a]D values, in conjunction with single-crystal X-ray diffraction analysis of the l-xylo stereoisomer. Examination of
the inhibitory properties of these imidazolo-pyrrolidinoses against six commonly encountered glycosidases led to the conclusion
that by and large the l-stereoisomers are inactive, whereas three out the four d-stereoisomers proved to be poor to moderate inhi-
bitors. It appears therefore that the most basic N(1) atom is not located in an optimal topology to be protonated easily inside the
enzyme’s active site.
# 2003 Elsevier Ltd. All rights reserved.
Introduction
synthesised the de-branched and at C(8) hydroxylated
analogue 2 of nagstatine as well as a series of stereomers
of 2.5 The inhibitory properties of 2 proved to be very
pronounced indeed with b-d-galactosidase (Escherichia
coli): Ki=2 nM!6 More recently, we published the
synthesis of the l-arabino analogue azasugar 3 which
showed a Ki=1 mM for the same b-galactosidase.7 These
two Ki values of manmade imidazolo-sugars 2 and 3
proved without any ambiguity: (i) the good binding
properties of the (protonated) imidazole moiety, the
basic N(1) atom occupying the pseudoanomeric posi-
tion which leads to (lateral) protonation;8,9 (ii) the
importance of a well defined configuration; (iii) the
importance of the hydroxymethyl group for an optimal
binding (docking) of the imidazole-sugar into the en-
zyme’s active site.
Both retaining and inverting polysaccharide glycosi-
dases are thought to lead to transition states (TS) with a
pronounced oxocarbenium character during the
‘induced fit’ step.1,2 Since most natural oligo- and poly-
saccharides are chair conformed pyranoses, the ensuing
oxocarbenium type TS’s appear as flattened, that is half-
chair, conformations. In 1992, Aoyagi, Aoyama and
their collaborators published the structure of the natural
product nagstatine 1, and showed this imidazole-sugar to
be a very potent inhibitor of some glucosaminidases,
with a Ki=4 nM for the bovine N-acetyl-b-d-glucosami-
nidase kidney enzyme.3,4 The flattened half-chair con-
formation of the piperidinose ring of 1 is worth to be
noticed: once protonated by the enzyme at the site of its
most basic N(1) atom, (the positively charged) nag-
statine mimics rather well the above postulated cyclic
oxocarbenium type TS. In 1995, Tatsuta and his group
In 1988, the natural product nectrisine 4 had been iso-
lated as an immunomodulator from the culture broth of
the fungus Nectria lucida10 and further shown to be a
powerful inhibitor of yeast a-glucosidase.11 The some-
what flattened conformation of nectrisine 4—as com-
pared to the envelope conformation of a saturated
*Corresponding author. Tel.: +33-389-336871; fax: +33-389-336875;
e-mail: j.streith@uha.fr
0968-0896/03/$ - see front matter # 2003 Elsevier Ltd. All rights reserved.
doi:10.1016/S0968-0896(03)00402-4

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3560
pyrrolidinose—once again could be considered as
mimicking the TS geometry of an oligo- or poly-
saccharide in the enzyme’s active site. We surmised that
by replacing the imine double bond of 4 by an imidazole
moiety, we should be able to obtain potent glycosidase
inhibitors by analogy with the manmade nagstatine ser-
ies, albeit to the best of our knowledge no naturally
occurring imidazolo-pyrrolidinoses have been isolated
so far.
In a first approach, we turned our attention to type 5
imidazolo-pyrrolidinoses (actually, their carbohydrate
rings are pyrrolines)12—all eight stereomers have been
synthesised—13 in the hope that some of them would
show inhibitory activity of glycosidases, albeit the most
basic N(2) atom is located one bond farther away with
respect to the N(1) atom of the reference model com-
pound 1. It turned out that the d-arabino, d-xylo, and
d-lyxo stereoisomers did show some activity, the d-ara-
bino stereoisomer 5 being the strongest inhibitor
(Ki=5 mM with a-d-mannosidase of Jackbeans). On the
other hand, the d-ribo and the four l stereoisomers in
the type 5 series proved to be either inactive or only
weakly active, at least with the six glycosidases we tested
(Fig. 1).13
Figure 2. The eight imidazolo-[1,2]-pyrrolidinoses.
Results and Discussion
Retrosynthetic analysis (Scheme 1)
Taking but one sequence to illustrate our approach, the
retrosynthetic analysis of both d-arabino and d-ribo
imidazolo-pyrrolidino-pentoses from an l-threose pre-
cursor is represented in Scheme 1. We had already
shown that nucleophilic addition of a C(2)-lithiated
imidazole to a protected l-threose led to a mixture of
the crystalline linear l-xylo and l-lyxo derivatives.7 We
expected the intramolecular cyclisation of these latter
two compounds—properly protected and activated—to
give the d-arabino and the d-ribo target molecules,
respectively. As will be described below, these retro-
synthetic assumptions could be translated into reality.
Like in the type 2 and type 3 piperidinose series, it
seemed of interest to have the basic N atom located in
the pseudoanomeric N(1) position in the pyrrolidinose
series, rather than one bond farther away. These con-
siderations led us to synthesise all eight N(1) type 6
imidazolo-pyrrolidinoses (Fig. 2) and to determine their
inhibitory properties with a series of six glycosidases.
The experimental results we obtained along these lines
are described and discussed below.
Arabinose and ribose series (Scheme 2)
D-Series. Reaction of the known l-xylo derivative 107
with TBDPSCl in the presence of Et3N and catalytic
amounts of DMAP in methylene chloride led to selec-
tive silylation of the primary alcohol, affording thereby
compound 11. Reaction of triflic anhydride with 11 in
the presence of pyridine at low temperature gave at once
bicyclic compound 14, obviously via a Walden inversion
of the short-lived triflic ester intermediate of 11. That
latter triflate, being too active, could not be isolated.
Treatment of 14 with fluoride ion (TBAF in THF) gave
15. Hydrogenolysis (H2/Pd/C) of the two O-benzyl
bonds of 15 led to the expected d-arabino target molecule
6. The same reaction conditions when applied to the
known l-lyxo derivative 12 led to d-ribo diastereomer 7,
Scheme 1. Retrosynthetic pathway for both the d-arabino- and
d-ribo-imidazolo-[1,2]-pyrrolidinoses.
Figure 1. Various imidazolosugar inhibitors of glycosidases.

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3561
Xylose and lyxose series (Scheme 3)
The synthetic schemes for the xylo and lyxo series
parallel those we have described above for the arabino
and ribo series (for the details, see Experimental).
D-Series. Reaction of the known l-arabino derivative
187 with TBDPS gave 19 whose triflate spontaneously
cyclised to give bicyclic compound 22. Sequential
removal of the three protecting groups gave the target
d-xylo imidazolo-sugar 8, the mono-alcohol di-O-ben-
zyl derivative 23 being the result of the first step. A
similar sequence of reactions starting from the known
l-ribo derivative 20 gave the d-lyxo imidazolo-sugar 9,
sequentially via intermediate 21, the short-lived triflate
of 21, and thence via intermediates 24 and 25.
L-Series. Reaction of the known d-arabino derivative
ent-18,7 again with TBDPS, gave ent-19 whose cyclisa-
tion led to ent-22. Removal of the protecting groups of
the latter compound gave the target l-xylo imidazolo-
sugar ent-8. A similar reaction sequence starting from
the known d-ribo derivative ent-207 gave the l-lyxo
imidazolo-sugar ent-9 (see Experimental).
Scheme 2. Reagents and conditions for the synthesis of the d-arabino-
and d-ribo-imidazolo-[1,2]-pyrrolidinoses and their enantiomers:
(a) CH2Cl2 or DMF, NEt3, TBDPSCl, DMAP cat; (b) CH2Cl2, pyri-
dine, Tf2O, ꢀ15 ꢁC to rt; (c) THF, TBAF; (d) Pd catalyst, H2 gas.
Spectral properties and structure analysis
sequentially via compound 13, the short-lived triflate of
13, thence via intermediates 16 and 17.
Structures and absolute configuration of the eight
stereisomers 6–9 and ent-6-ent-9 (Fig. 2) could be
determined unambiguously, thanks to a combination of
1H/13C NMR and chiroptical data analyses. These
structural assignments are also due to the fact that the
absolute 3-D structures of the corresponding eight lin-
ear imidazolo-pentose precursors (i.e., 10/ent-10, 12/ent-
12, 18/ent-18, and 20/ent-20) had been determined pre-
viously without any ambiguity, thanks to a combination
L-Series. The syntheses of the ent-6 and ent-7 enantio-
mers were performed using the same sequence of
reactions as in Scheme 2, in the enantiomeric series
though. In other words, the known enantiomers ent-10
and ent-117 were the starting materials for the syn-
thesis of the l-arabino ent-6 and l-ribo ent-7 enantio-
mers, respectively.
The experimental results showed the final target mole-
cules to have the expected physical properties. In parti-
cular, the chiroptical data clearly demonstrated the
mirror-image relationship between 6 and ent-6, and
between 7 and ent-7 (see Table 1 for [a]D values, and the
Experimental for CD spectra).
20
Table 1. ½aŠ values measured in MeOH (c=1)
D
+23
+57
+51
+36
Scheme 3. Reagents and conditions for the synthesis of the d-xylo-
and d-lyxo-imidazolo-[1,2]-pyrrolidinoses and their enantiomers: (a)
CH2Cl2 or DMF, NEt3, TBDPSCl, DMAP cat; (b) CH2Cl2, pyridine,
Tf2O, ꢀ15 ꢁC to rt; (c) THF, TBAF; (d) Pd catalyst, H2 gas.
ꢀ23
ꢀ53
ꢀ55
ꢀ35

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3562
1
of H/13C NMR spectroscopy, circular dichroism (CD)
and rotatory power data, in conjunction with two sin-
gle-crystal X-ray diffraction analyses.7 Furthermore
single crystal X-ray diffraction analysis of ent-8 con-
firmed its l-xylo configuration, proving thereby the
Walden inversion of its d-arabino precursor (Fig. 3).
During the key reaction step—that is, the intramolec-
ular cyclisation—a Walden inversion occurs which
brings about a configurational change from the d- to
the l-series, or vice versa from the l- to the d-series.
For example, the linear l-arabino precursor 18 leads to
the bicyclic imidazolo d-xylo target azasugar 8 (Scheme
3); which in the enantiomeric series reads as follows: the
linear d-arabino precursor ent-18 leads to the bicyclic
imidazolo l-xylo target azasugar ent-8.
by the absolute configuration of carbon atom C(5), a
(5R) configuration leading to a positive sign, a (5S)
configuration to a negative one. In a previous publica-
tion we observed similar chiroptical properties with all
type 5 stereomers [in these isomers the basic N(2) atom
is one bond farther away with respect to type 6 isomers]:
type 5 d-stereomers proved to be dextroratatory, their
l-enantiomers levorotatory.13
As to the CD spectra of the eight type 6 target azasu-
gars, they are definite testimony to the mirror image
relationship between pairs of opposite enantiomers
6/ent-6, 7/ent-7, 8/ent-8 and 9/ent-9 (see Experimental).
Enzymatic assays
Chiroptical properties proved to be of great help and
interest. The optical rotatory power data agree rather
well with the mirror image relationship between opposite
enantiomers (Table 1). In terms of [a]D values, it is worth
noting that all d-configured imidazolosugars are dex-
trorotatory, the l-stereomers being levorotatory. The
sign of these [a]D values seems to be largely determined
The eight imidazolo-pyrrolidino-pentoses (Fig. 2) have
been evaluated as potential inhibitors of six commer-
cially available glycosidases, and the results compiled in
Table 2. In the d-series, those imidazolo sugars which
do show some activity proved to be competitive inhibi-
tors, but only moderate inhibitions could be measured.
Both the d-arabino 6 and the d-ribo 7 stereomers inhibit
a-d-mannosidase (Jackbeans), with Ki values of 36 and
90 mM, respectively. Notice that in stereomer 6 all three
configurations C(7), C(6) and C(5) are identical with
respect to those of the corresponding asymmetric cen-
tres C(3), C(4) and C(5) of d-mannose. d-Stereomer 6
inhibits also a-d-glucosidase (baker’s yeast), though
only poorly (Ki=160 mM), albeit in that instance too we
notice a perfect configurational fit with respect to the
asymmetric centres C(3), C(4) and C(5) of d-glucose.
The four stereomers of the l-series are essentially inac-
tive, at least with the six glycosidases which were tested.
It appears therefore that the eight imidazolo-pyrrazolo-
pentoses of Figure 2 are poor inhibitors, if at all, of hex-
ose-glycosidases. Obviously, the most basic N(1) atom is
not located in an optimal topology—to be protonated
easily in the enzyme’s active site—whatever the degree of
induced fit, or stated differently, whatever the degree of
configurational complementarity inside the old ‘lockand
key’ model. One may also reach the conclusion that the
above postulated structural analogy between nectrisine
and the eight azasugars of Figure 2 is a bit far-fetched,
the most basic N(1) atom of the herein described imi-
dazolosugars being located in quite a different position
when compared to the one of nectrisine’s N atom.
Figure 3. ORTEP plot of the structure of ent-8 (50% probability
ellipsoıds).
Table 2. Inhibition values of the imidazolo-[1,2]-pyrrolidinoses against six commercially available glycosidases
a-d-Glucosidase (baker’s yeast)
b-d-Glucosidase (almonds)
Ki=160 mM
NI
NI
NI
NI
NI
Ki=64 mM
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
NI
Ki=100 mM
a-d-Galactosidase (green coffee beans)
b-d-Galactosidase (Escherichia coli)
a-d-Mannosidase (Jackbeans)
NI
NI
NI
NI
NI
NI
NI
NI
60% inhibition
NI
Ki=36 mM Ki=90 mM
NI NI
b-d-Mannosidase (acetone snail powder)

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3563
Experimental
addition of enzyme in a 1-mL assay medium (acetate
buffer 50 mM, or phosphate buffer 20 mM according to
the desired pH value) using substrate concentrations
around the Km value of each enzyme. The Ki values were
determined for the most potent inhibitors, by the Dixon
graphical procedure.15,16
General
Flash chromatography (FC): silica gel (Merck60;
230–400 mesh). TLC: silica gel on aluminium sheets
(Merck60HF 254); the spots were viewed under UV or
by heating with a thermogun after spraying with a
solution of KMnO4 (20 g) and Na2CO3 (40 g) in H2O
(1 L) or a solution of phosphomolybdic acid (5% in
96% EtOH). Mp: Kofler hot-bench or Buchi-SMP
apparatus; corrected values. Optical rotations were
measured at +20 ꢁC: Schmidt-Haensch Polartronic
Universal polarimeter. CD spectra were measured in
H2O solution between 180 and 400 nm under nitrogen
with a Jobin Yvon CD6 Dichrograph (ꢀe values) at the
research centre of the Roche pharmaceutical division in
1,2-di-O-Benzyl-4-O-TBDPS L-xylo derivative 11. What
follows shallꢁbe called Procedure A: A solution of 10,
mp=99–100 C7 (214 mg, 0.58 mmol), DMAP (7 mg,
60 mmol)), anhydrous Et3N (100 mL, 0.7 mmol) and
TBDPSCl (166 mL, 0.64 mmol) in anhydrous CH2Cl2
(5 mL) was stirred at room temp. After 48 h, the solu-
tion was diluted with CH2Cl2 (20 mL), and washed with
a saturated solution of NH4Cl (20 mL). The aqueous
solution was separated, extracted with CH2Cl2 (2 Â
30 mL), and the combined organic fractions were dried
(MgSO4), filtered, and evaporated to dryness. The resi-
due was purified by chromatography (AcOEt/cyclohex-
ane 1:1) to give 11 (186 mg, 53%) as a colourless foam.
1H NMR (CDCl3, 400 MHz): d=0.95 [s, 9H, (CH3)3C],
3.58 [d, 2H, C(4)–Ha and C(4)–Hb], 3.80 [td, 1H, C(3)–
H], 3.87 [dd, 1H, C(2)–H], 4.28 and 4.37 [AB, 2H,
J=11.6, OCH2Ph], 4.44 and 4.48 [AB, 2H, J=11.2,
OCH2Ph], 4.87 [d, 1H, C(1)–H], 6.91 [slarge, 1H, C(20)–H
or C(30)–H], 7.06 (slarge, 1H, C(30)–H or C(20)–H], 7.10–
7.33 and 7.49–7.57 [m, 20H, H–arom phenyl], J1,2=6.0,
J2,3=2.4, J3,4=6.4. 13C NMR (CDCl3, 100.6 MHz):
d=19.0 [SiC(CH3)3], 26.8 [SiC(CH3)3], 64.3 [C(4)], 70.8
[C(3)], 71.7 [CH2Ph], 74.4 [CH2Ph], 76.5 [C(1)], 79.3
[C(2)], 126.9 [large hump, C(20) and C(30)], 127.6–128.2
(C–arom phenyl)], 129.6, 133.0 and 133.1 [Cs phenyl)],
135.39–135.41 [C–arom phenyl], 137.4 and 137.9 [Cs
phenyl], 145.5 [C(20)]
Basel, Switzerland. H and 13C NMR spectra: 250 and
1
62.9 MHz, respectively (Bruker ACF-250 spectrometer
at 300 K) or 400 and 100.6 MHz, respectively (Bruker
DSX-400 spectrometer at 300 K). Internal references for
1H NMR: SiMe4 (d=0.00), CDCl3 (d=7.26), CD3OD
(d=3.30), [D4]TSP for spectra in D2O (d=0.00); for 13
C
NMR: CDCl3 (d=77.03), CD3OD (d=49.02); d in ppm
and J in Hz. HR-MS were measured with ESI mode in
the departments of spectroscopy of Novartis in Basle
and of the Faculte de Chimie, Universite Louis Pasteur
at Strasbourg. Microanalyses were carried out by the
Service Central de Microanalyses of the CNRS, 69390
Vernaison, France. ‘MeOH+NH3’ stands for a solution
of pure MeOH saturated at room temp with NH3 (ex
gas form). Four different recipes were used, each one
eight times. Each recipe is described, only once in full
detail, along with its workup and chromatographic
methodologies, the entire procedure being called either
Procedure A, Procedure B, Procedure C, or Procedure D.
In some instances, minor modifications were used; they
are indicated explicitly.
1,2-di-O-Benzyl-4-O-TBDPS D-xylo derivative ent-11.
Procedure A as described above, starting from ent-10,
mp=103–104 ꢁC7 (200 mg, 0.54 mmol), DMAP (ca.
6 mg), anhydrous Et3N (114 mL), in DMF (7 mL), and
TBDPSCl (114 mL, 0.65 mmol). After 48 h, workup as
above using CH2Cl2 (30 mL), NH4Cl (30 mL), (MgSO4),
and chromatography to give ent-11 (177 mg, 54%) as a
Enzymatic assays
Glycosidases [a-mannosidase (EC 3.2.1.24) from Jack
beans (Sigma M 7257), b-mannosidase (EC 3.2.1.25)
from snail acetone powder (Sigma M 9400), a-glucosi-
dase (EC 3.2.1.20) from baker’s yeast (Sigma G-5003),
b-glucosidase (EC 3.2.1.21) from almonds (Sigma
G-4511), a-galactosidase (EC 3.2.1.22) from green cof-
fee beans (Sigma G-8507), b-galactosidase (EC 3.2.1.23)
from E. coli (Sigma G-4155)], and their corresponding
substrates were purchased from Sigma Co. Spectro-
photometric assays were performed at the optimum pH
for each enzyme,14 with p-nitrophenyl-a-d-mannopyr-
anoside as a substrate for a-mannosidase (Km=2 mM,
pH=4.5), p-nitrophenyl-b-d-mannopyranoside for
b-mannosidase (Km=1.33 mM, pH=4.0), p-nitrophe-
nyl-a-d-glucopyranoside for a-glucosidase (Km=0.3
mM, pH=7), p-nitrophenyl-b-d-glucopyranoside for b-
glucosidase (Km=1.3 mM, pH=5.0), p-nitrophenyl-a-
d-galactopyranoside for a-d-galactosidase (Km=0.25
mM, pH=6.5) and p-nitrophenyl-b-d-galactopyrano-
side for b-d-galactosidase (Km=0.4 mM, pH=7). The
release of p-nitrophenol was measured continuously at
405 nm to determine initial velocities. All kinetics were
performed at 25 ꢁC and the reaction was started by the
1
colourless foam. H (CDCl3, 400 MHz) and 13C NMR
(CDCl3, 100.6 MHz) spectra of ent-11 proved to be
identical to, and superimposable on, those of the l-xylo
enantiomer 11.
1,2-di-O-Benzyl-4-O-TBDPS L-lyxo derivative 13. Pro-
cedure A starting from 12, mpdec=161 ꢁC7 (500 mg,
1.36 mmol), DMAP (ca. 7 mg, 60 mmol), anhydrous
Et3N (245 mL, 1.77 mmol), TBDPSCl (425 mL,
1.63 mmol) in anhydrous CH2Cl2 (15 mL). After 24 h
workup and purification by chromatography as above
to give 13 (719 mg, 87%) as a colourless foam. 1H NMR
(CDCl3, 250 MHz): d=1.05 [s, 9H, (CH3)3C], 3.69 [m,
3H, C(3)-H, C(4)-Ha and C(4)-Hb], 4.23 [t, 1H, C(2)-H],
4.51 [s, 2H, OCH2Ph], 4.60 and 4.70 [AB, 2H, J=11.0,
OCH2Ph], 4.81 [d, 1H, C(1)-H], 6.95 [slarge, 1H, C(40)-H
or C(50)-H], 7.05 (slarge, 1H, C(50)-H or C(40)-H],
7.23–7.65 [m, 22H, H-arom phenyl], 9.54 [slarge, 1H,
N–H]; J1,2=3.7, J2,3=3.7 (n.b.: the other J values could
not be determined, due to the fact that C(3)-H, C(4)-Ha
and C(4)-Hb appear as a complex multiplet).

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3564
1,2-di-O-Benzyl-4-O-TBDPS D-lyxo derivative ent-13.
Procedure A, starting from ent-12, mpdec=161 ꢁC7
(400 mg, 1.08 mmol), DMAP (catalytic amount), anhy-
drous Et3N (227 mL, 1.63 mmol), TBDPSCl (229 mL,
1.30 mmol) in anhydrous CH2Cl2 (15 mL). After 24 h at
rt, workup and chromatography led to ent-13 (564 mg,
133.7 [C(2)], 137.1 and 137.4 [Cs phenyl], 150.7 [C(7a)].
HR-MS: [M+H]+ ion 351.1703 (C21H23N2O3, calcd
351.1709).
6,7-di-O-Benzyl-8-OTBDPS L-arabino derivative ent-14
and 6,7-di-O-benzyl L-arabino derivative ent-15. Proce-
dure B starting from ent-11 (113 mg, 0.19 mmol), and
anhyd pyridine (45 mM, 0.56 mmol) in anhyd CH2Cl2
(3 mL) at ꢀ15 ꢁC to which Tf2O (77 mL, 0.46 mmol) was
added. After 12 h at rt workup and purification by
chromatography gave ent-14 (88 mg, 80%) as a colour-
87%) as
a
colourless foam. 1H NMR (CDCl3,
250 MHz) spectrum was identical to, and super-
imposable on, the one of its enantiomer 13.
6,7-di-O-Benzyl-8-OTBDPS D-arabino derivative 14 and
6,7-di-O-benzyl D-arabino derivative 15. What follows
shall be called Procedure B for the preparation of 14,
and Procedure C for the preparation of 15.
1
less foam which was used for the next reaction step. H
NMR (CDCl3, 400 MHz): d=0.97 [s, 9H, (CH3)3C],
3.73 [dd, 1H, C(8)-Hb], 3.80 [dd, 1H, C(8)-Ha], 4.14
[ddd, 1H, C(5)-H], 4.26 [dd, 1H, C(6)-H], 4.42 and 4.51
[AB, 2H, J=11.7, OCH2Ph], 4.71 and 4.93 [AB, 2H,
J=11.7, OCH2Ph], 4.76 [d, 1H, C(7)-H], 6.91 [d, 1H,
C(3)-H], 7.09 [d, 1H, C(2)-H], 7.12–7.55 [m, 20H,
H-arom phenyl], J2,3=1.1, JHa,Hb=10.8, JHa,5=4.8,
JHb,5=7.6, J5,6=3.3, J6,7=2.4. That 1H NMR spectrum
matched the one of compound 14. 13C NMR (CDCl3,
100.6 MHz): d=19.14 [SiC(CH3)3], 26.75 [SiC(CH3)3],
63.8 [C(5)], 64.6 [C(8)], 71.2 [OCH2Ph], 71.9 [OCH2Ph],
77.0 [C(7)], 87.9 [C(6)], 114.9 [C(3)], 127.6-128.5
[C–arom phenyl], 129.7 [C–arom phenyl], 130.0 [C–arom
phenyl], 132.7 and 132.6 [Cs phenyl], 133.4 [C(2)], 135.5
and 137.6 [C–arom phenyl], 137.1 and 137.7 [Cs phenyl],
150.6 [C(7a)].
Procedure B. To a stirred solution of 11 (180 mg,
0.30 mmol) and anhydrous pyridine (95 mL, 1.2 mmol) in
anhydrous CH2Cl2 (5 mL) at ꢀ15 ꢁC under argon
atmosphere was added dropwise Tf2O (150 mL,
0.90 mmol). The reaction mixture was stirred at ꢀ15 ꢁC
for 15 min, and finally 12 h at rt. The resulting mixture
was diluted with CH2Cl2 (30 mL), and the organic phase
washed with a saturated solution of NH4Cl (30 mL).
The aqueous phase was separated and extracted with
CH2Cl2 (2 Â 20 mL). The combined organic fractions
were dried (MgSO4), filtered and evaporated to dryness.
The residue was purified by chromatagraphy (AcOEt/
cyclohexane 1:1) to give 14 (93 mg, 53%) as a colourless
foam which was used as such for the next reaction step.
1H NMR (CDCl3, 250 MHz): d=0.97 [s, 9H, (CH3)3C],
3.73 [dd, 1H, C(8)-Hb], 3.81 [dd, 1H, C(8)-Ha], 4.14
[ddd, 1H, C(5)-H], 4.26 [dd, 1H, C(6)-H], 4.42 and 4.51
(AB, 2H, J=11.6, OCH2Ph], 4.71 and 4.93 [AB, 2H,
J=11.6, OCH2Ph], 4.76 [d, 1H, C(7)-H], 6.90 [d, 1H,
C(3)–H], 7.08 [d, 1H, C(2)-H], 7.11–7.60 [m, 20H,
H–arom phenyl], J1,2=1.2, JHa,Hb=10.7, JHa,5=4.6,
JHb,5=7.3, J5,6=3.4, J6,7=2.1.
Procedure C. A solution of ent-14 (144 mg, 0.24 mmol)
in anhydrous THF (3.8 mL) was treated with a 1 M
solution of TBAF in THF (370 mL, 0.37 mmol) as
above. Workup and purification with chromatography
as above led to ent-15 (83 mg, 97%) as a colourless oil.
½aŠ2D0=+35 (c 2, CHCl3). 1H NMR (CDCl3), 400 MHz):
d=3.70 [dd, 1H, C(8)-Hb], 3.84 [dd, 1H, C(8)-Ha], 4.15
[ddd, 1H, C(5)-H], 4.28 [t, 1H, C(6)-H], 4.49 and 4.56
[AB, 2H, J=11.8, OCH2Ph], 4.61 and 4.89 [AB, 2H,
J=11.6, OCH2Ph], 4.74 [d, 1H, C(7)-H], 6.92 [d, 1H,
C(2)-H], 7.03 [d, 1H, C(3)-H], 7.18–7.30 [m, 10H,
H–arom phenyl], J2,3=1.1, JHa,Hb=11.6, J5,Ha=4.5,
J5,Hb=6.8, J5,6=2.4, J6,7=2.1. 13C NMR (CDCl3,
100.6 MHz): d=62.8 [C(8)], 64.0 [C(5)], 71.3 [OCH2Ph],
71.9 [OCH2Ph], 76.6 [C(7)], 88.1 [C(6)], 115.0 [C(3)],
127.8–128.5 [C–arom], 133.1 [C(2)], 137.0 and 137.3 [Cs
Procedure C. To a stirred solution of the preceding
compound 14 (136 mg, 0.23 mmol) in anhydrous THF
(3 mL) was added dropwise a 1 M solution of TBAF in
THF (600 mL; 0.58 mmol). The reaction mixture was
stirred at room temp for 2 h, concentrated to dryness,
the residue dissolved in CH2Cl2 (30 mL) and the result-
ing solution was washed with a saturated solution of
NH4Cl (40 mL). The aqueous phase was separated and
extracted with CH2Cl2 (2 Â 30 mL) and the combined
organic fractions were dried (MgSO4), filtered and eva-
porated to dryness. The resulting residue was purified
by chromatogaphy (Et2O/MeOH–NH3 97:3) to give 15
(65 mg, 43%) as a colourless oil. ½aŠ2D0=ꢀ40 (c 2,
1
phenyl], 150.35 [C(7a)]. These H and 13C NMR spectra
matched the ones of compound 15. HR-MS: [M+H]+
ion: 351.1715 (C21H23N2O3), calcd 351.1709.
6,7-di-O-Benzyl-8-OTBDPS D-ribo derivative 16 and
6,7-di-O-benzyl D-ribo derivative 17. Procedure B, start-
ing from 13 (710 mg, 1.17 mmol), anhyd pyridine
(370 mL) in anhyd CH2Cl2 (15 mL) at ꢀ15 ꢁC to which
Tf2O (590 mL, 3.51 mmol) was added dropwise. After
12 h at rt workup as above followed by chromatography
to give 16 as a colourless foam which was used as such
1
CHCl3). H NMR (CDCl3, 250 MHz): d=3.80 (dd, 1H,
C(8)-Hb], 3.93 [dd, 1H, C(8)-Ha], 4.24 [ddd, 1H, C(5)-
H], 4.37 [t, 1H, C(6)-H], 4.57 and 4.65 [AB, 2H, J=11.7,
OCH2Ph], 4.79 and 5.00 [AB, 2H, J=11.9, OCH2Ph],
4.83 [d, 1H, C(7)-H], 7.01 [d, 1H, C(3)-H], 7.15 [d, 1H,
C(2)-H], 7.27–7.42 [m, 10H, H–arom phenyl]. J2,3=1.1,
1
for the next step. H NMR (CDCl3, 250 MHz): d=0.98
[s, 9H, (CH3)3C], 3.82 [dd, 1H, C(8)-Hb], 4.05 [dd, 1H,
C(8)-Ha], 4.22 [dd, 1H, C(6)-H], 4.39 and 4.71 [AB, 2H,
J=11.7, OCH2Ph], 4.44 [ddd, 1H, C(5)-H], 4.71 and
4.86 [AB, 2H, J=11.9, OCH2Ph], 4.74 [d, 1H, C(7)-H],
7.00 [d, 1H, C(2)-H or C(3)-H], 7.16 [d, 1H, C(3)-H or
JHa,Hb=11.4,
J5,Ha=4.4,
J5,Hb=6.4,
J5,6=2.9,
J6,7=2.1. 13C NMR (CDCl3, 62.9 MHz): d=63.2 [C(8)],
63.9 [C(5)], 71.3 [OCH2Ph], 72.1 [OCH2Ph], 76.4 [C(7)],
88.3 [C(6)], 114.8 [C(3)], 127.9–128.6 [C–arom phenyl],

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3565
C(2)-H], 7.20–7.59 [m, 20H, H–arom phenyl], J2,3=1.0,
JHa,Hb=11.2,
J6,7=5.5.
MeOH. The combined organic solutions were evapo-
rated to dryness in vacuum, and the residue purified by
chromatography (AcOEt/MeOH 7:3) to give 6 after
lyophilisation (74 mg, 80%) as colourless hygroscopic
microcrystals. Mpdec=172–174 ꢁC. ½aŠD20=+23 (c 1,
MeOH). CD (H2O): recording starts at 180.0 (+8.27),
195 (0.00), 199.5 (ꢀ1.25), 205.0 (0.00), 210.5 (+0.68),
222 (0.00), 230 (ꢀ0.33), tailing out at ca. 245 (ꢀ0.10). 1H
NMR (D2O, 250 MHz): d=3.91 [dd, 1H, C(8)-Hb], 4.13
[dd, 1H, C(8)-Ha], 4.20 [q, 1H, C(5)-H], 4.51 [t, 1H, C(6)-
H], 4.95 [d, 1H, C(7)-H], 7.17 [sbroad, 1H, C(2)-H], 7.22 [s,
1H, C(3)-H], JHa,Hb=12.2, JHa,5=3.7, JHb,5=5.2,
J5,6=4.1, J6,7=3.9. 13C NMR (CD3OD, 62.9 MHz):
d=63.0 [C(8)], 66.8 [C(5)], 74.2 [C(7)], 84.0 [C(6)], 116.1
[C(3)] and 132.8 [C(2)], 153.4 [C(7a)]. HR-MS: [M+H]+
ion 171.0770 (C7H11N2O3, calcd 171.0770).
J5,Ha=2.5,
J5,Hb=5.9,
J5,6=7.1,
Procedure C. To a stirred solution of the preceding
product 16 in anhydrous THF (20 mL) was added
dropwise a 1 M solution of TBAF in THF (2 mL,
2.0 mmol). After 12 h at rt workup as above followed by
chromatography (Et2O/MeOH–NH3 97:3) gave 17
(340 mg, 83%) as colourless crystals. Mp=115–116 ꢁC.
½aŠ2D0=+202 (c 2, CHCl3). 1H NMR (CDCl3, 250 MHz):
d=2.90 [slarge, OH], 3.77 [dd, 1H, C(8)-Hb], 4.13 [dd,
1H, C(8)-Ha], 4.23 [dd, 1H, C(6)-H], 4.40 [ddd, 1H,
C(5)-H], 4.47 and 4.76 [AB, 2H, J=11.5, OCH2Ph], 4.65
and 4.82 [AB, 2H, J=11.8, OCH2Ph], 4.70 [d, 1H, C(7)-
H], 7.06 [d, 1H, C(2)-H or C(3)-H], 7.11 [d, 1H, C(3)-H
or C(2)-H], 7.28–7.45 [m, 10H, H–arom phenyl],
L-arabino-Imidazolo-pyrrolidinose ent-6. Procedure D,
starting from ent-15 (141 mg, 0.40 mmol), 20%
Pd(OH)2/C (+ 30% H2O) in EtOH (3 mL) and AcOH
(3 mL) under H2 pressure (ca. 1.5 bar). After 16 h
workup as above. The clear organic solution was con-
centrated in vacuum and AcOH was almost entirely
removed via three azeotropic distillations with toluene.
Last traces of AcOH were removed with basified IRA
400 (OHꢀ) resin. The crude residue was purified by
chromatography (CH2Cl2/MeOH 8:2) to give ent-6
(65 mg, 95%) as a colourless solid. Mpdec=175–176 ꢁC
J2,3=1.1,
JHa,Hb=11.8,
J5,Ha=2.8,
J5,Hb=5.8,
J5,6=7.1, J6,7=5.4. 13C NMR (CDCl3, 62.9): d=61.6
[C(8)], 62.0 [C(5)], 69.1 [C(7)], 70.3 [OCH2Ph], 72.2
[OCH2Ph], 80.4 [C(6)], 114.5 [C(3)], 127.8–128.5
[C–arom phenyl], 132.6 [C(2)], 137.1 and 137.6 [Cs
phenyl], 150.2 [C(7a)]. C21H22N2O3 (350.4): C 71.98, H
6.33, N 7.99; found: C 71.7, H 6.3, N 7.9.
6,7-di-O-Benzyl-8-OTBDPS L-ribo derivative ent-16 and
6,7-di-O-benzyl L-ribo derivative ent-17. Preparation of
ent-16. Procedure B, starting from ent-13 (546 mg,
0.90 mmol), anhydrous pyridine (218 mL) in anhydrous
CH2Cl2 (15 mL) and Tf2O (367 mL). After 12 h at rt
workup followed by chromatogaphy gave ent-16
(389 mg, 73%) as a colourless foam which was used as
such in the following step. 1H NMR (CDCl3, 250 MHz):
spectrum identical to, and superimposable on, that of
16. 13C NMR (CDCl3, 62.9 MHz): d=19.0 [SiC(CH3)3],
26.5–26.8 [SiC(CH3)3], 62.0 [C(8)], 62.7 [C(5)], 68.8
[C(7)], 70.1 [OCH2Ph], 71.9 [OCH2Ph], 80.0 [C(6)],
114.6 [C(3)], 127.7–129.9 [C–arom phenyl], 132.3 and
132.4 [Cs phenyl], 132.7 [C(2)], 135.4–135.5 [C–arom
phenyl], 137.0 and 137.5 [Cs phenyl], 150.0 [C(7a)].
(MeOH). ½aŠ20=ꢀ23 (c 1, MeOH). CD (H2O): record-
D
ing starts at 180.0 (ꢀ8.70), 182 (ꢀ8.83), 195 (0.00), 200.5
(+0.80), 205 (0.00), 210 (ꢀ1.07), 222 (0.00), 229
(+0.20), tailing out at ca. 248 (0.0). 1H NMR (CD3OD,
400 MHz): d=3.75 [dd, 1H, C(8)-Hb], 4.00 [m, 2H,
C(8)-Ha and C(5)-H], 4.31 [t, 1H, C(6)-H], 4.72 [d, 1H,
C(7)-H], 7.02 [d, 1H, C(2)-H], 7.12 [d, 1H, C(3)-H];
J2,3=1.3,
JHa,Hb=11.3,
JHa,5=4.0,
JHb,5=6.1,
J5,6=3.5, J6,7=3.3. 13C NMR (CD3OD, 100.6 MHz):
d=63.0 [C(8)], 66.9 [C(5)], 74.2 [C(7)], 84.1 [C(6)], 116.1
[C(3)], 132.8 [C(2)], 153.5 [C(7a)]. These two NMR
spectra matched those of enantiomer 6. HR-MS:
[M+H]+ ion 171.0768 (C7H11N2O3, calcd 171.0770).
D-ribo-Imidazolo-pyrrolidinose 7. Procedure D. A stirred
solution of 17 (164 mg, 0.47 mmol) in AcOH (5 mL)
containing 20% Pd(OH)2/C (+ 30% H2O)(150 mg) was
put under H2 pressure (20 bar) at rt. After 2 days,
workup as above and purification of the crude residue
by chromatography (AcOEt/MeOH 7:3) which gave 7
(61 mg, 76%) as a colourless oil which was put in H2O
solution and deep-freezed. After lyophilisation it led to
7 (61 mg, 76%) a colourless powder which turned
slightly pink. Mpdec=105–107 ꢁC. ½aŠD20=+57 (c 1,
MeOH). CD (H2O): recording starts at 180.00 (ꢀ0.71),
184.5 (ꢀ4.40), 195 (0.00), 205.5 (+2.11), tailing out at
Preparation of ent-17. Procedure C, starting from ent-
16 (385 mg, 0.65 mmol) in anhydrous THF (10 mL),
containing a 1 M solution of TBAF in THF (0.98 mL,
0.98 mmol). After 5 h at rt, workup as above, followed
by chromatography gave ent-17 (220 mg, 96%) as a
colourless solid. Mp=116–116.5 ꢁC. ½aŠD20=ꢀ192 (c=2,
1
CHCl3). H NMR (CDCl3, 250 MHz) and 13C NMR
(CDCl3, 62.9 MHz) spectra identical to, and super-
imposable on, those of compound 17. C21H22O3N2
(350.4): C 71.98, H 6.33, N 7.99; found: C 72.2, H 6.3, N
8.1.
1
ca. 246 (ꢀ0.15). H NMR (CD3OD, 400 MHz): d=3.77
D-arabino-Imidazolo-pyrrolidinose 6. What follows shall
be called Procedure D: A stirred solution of 15 (190 mg,
0.54 mmol) in MeOH (5 mL) was put under H2 pressure
(30 bar) in the presence of 20% Pd(OH)2/C (+30%
H2O) at rt, the reaction being monitored by TLC
(CH2Cl2/MeOH–NH3 2:1). After 7 days, the conversion
seemed to be complete. The suspension was centrifuged
and the catalyst was rinsed several times with hot
(dd, 1H, C(8)-Hb], 4.08 [dd, 1H, C(8)-Ha], 4.15 [td, 1H,
C(5)-H], 4.43 [t, 1H, C(6)-H], 4.82 [d, 1H, C(7)-H], 7.03
[d, 1H, C(2)-H], 7.14 [d, 1H, C(3)-H], JHa,Hb=11.9,
JHa,5=2.9, JHb,5=5.8, J5,6=5.6, J6,7=5.8, J2,3=1.0.
13C NMR (CD3OD, 100.6 MHz): d=62.2 [C(8)], 65.7
[C(5)], 66.9 [C(7)],76.5 [C(6)], 115.9 [C(3)], 132.4 [C(2)],
153.6 [C(7a)]. HR-MS: [M+H]+ ion 171.0770
(C7H11N2O3, calcd 171.0770).

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3566
L-ribo-Imidazolo-pyrrolidinose ent-7. Procedure D, start-
ing from a solution of ent-17 (271 mg, 0.77 mmol), in
EtOH (6 mL) and AcOH (6 mL) containing 20%
Pd(OH)2/C (+50% H2O) under H2 pressure (ca.
1.5 bar) at rt. After 12 h, workup and purification via
chromatography as above gave ent-7 (92 mg, 70%) as a
mp=109–110 ꢁC7 (530 mg, 1.44 mmol), imidazole
(140 mg), DMAP (10 mg), Et3N (265 mL) and
TBDPSiCl (454 mL, 1.73 mmol) in CH2Cl2 (16 mL).
After 48 h, workup and purification by chromatography
led to ent-21 (647 mg, 74%) as a colourless foam. H
NMR (CD3OD, 250 MHz) spectrum identical to, and
superimposable on, that of 21.
1
colourless oil. ½aŠ20=ꢀ53 (c 0.55, MeOH). CD (H2O):
D
recording starts at 180.0 (ca. 0.0), 185.5 (+2.40), 195
1
(0.00), 206.5 (ꢀ1.61), tailing out at ca. 245 (+0.05). H
6,7-di-O-Benzyl-8-OTBDPS D-xylo derivative 22 and
6,7-di-O-benzyl D-xylo derivative 23. Procedure B. To a
stirred solution of 19 (399 mg, 0.66 mmol) and anhy-
drous pyridine (210 mL, 2.64 mmol) in anhydrous
CH2Cl2 (10 mL) at ꢀ15 ꢁC under argon atmosphere was
added dropwise Tf2O (330 mL, 1.98 mmol). The reac-
tion mixture was stirred for 30 min at 0 ꢁC, then over-
night at rt. Workup as for 14, but without
chromatographic purification, led to crude compound
22 (477 mg) as an orange solid. 1H NMR (CDCl3,
250 MHz): d=0.96 [s, 9H, (CH3)3C], 3.81 [dd, 1H, C(8)-
Hb], 3.93 [dd, 1H, C(8)-Ha], 4.50 [ddd, 1H, C(5)-H], 4.57
and 4.72 [AB, 2H, J=11.7, OCH2Ph], 4.65 [dd, 1H,
C(6)-H], 4.93 and 5.15 [AB, 2H, J=11.8, OCH2Ph], 5.03
[d, 1H, C(7)-H], 7.02 [d, 1H, C(2)-H or C(3)-H],
7.23–7.61 [m, 21H, H–arom phenyl, C(2)-H or C(3)-H];
NMR (CD3OD, 400 MHz): d=3.77 [dd, 1H, C(8)-Hb],
4.07 [dd, 1H, C(8)-Ha], 4.14 [td, 1H, C(5)-H], 4.42 [t,
1H, C(6)-H], 4.82 [d, 1H, C(7)-H], 7.02 [d, 1H, C(2)-H],
7.14 [d, 1H, C(3)-H], JHa,Hb=11.8, JHa,5=2.8,
JHb,5=5.8, J5,6=5.7, J6,7=5.5, J2,3=1.3. 13C NMR
(CD3OD, 100.6 MHz): d=62.1 [C(8)], 65.8 [C(5)], 67.0
[C(7)], 76.4 [C(6)], 116.0 [C(3)] 132.2 [C(2)], 153.3
[C(7a)]. These two NMR spectra proved to be identical
to, and superimposable on, those of 7. HR-MS:
[M+H]+ ion 171.0768 (C7H11N2O3, calcd 171.0770).
1,2-di-O-Benzyl-4-O-TBDPS L-arabino derivative 19.
Procedure A, starting from
a
solution of 18,
mp=111–113 ꢁC7 (350 mg, 0.95 mmol), DMAP (7 mg,
60 mmol), anhydrous Et3N (170 mL, 1.0 mmol), and
TBDPSCl (295 mL, 1.14 mmol) in anhydrous CH2Cl2
(10 mL). Workup and chromatographic purification led
J2,3=1.0, JHa,Hb=10.9, JHa,5=5.4,
J5,6=6.8, J6,7=4.3.
J
Hb,5=3.2,
1
to 19 (405 mg, 70%) as a colourless foam. H NMR
(CDCl3, 250 MHz): d=1.07 [s, 9H, (CH3)3C], 2.87 [d,
1H, OH], 3.77 [dd, 1H, C(4)-Hb], 3.82 [dd, 1H, C(4)-Ha],
3.84 [dd, 1H, C(2)-H], 3.93 and 4.27 [AB, 2H, J=10.8,
OCH2Ph], 3.96 [m, 1H, C(3)-H], 4.47 [s, 2H, OCH2Ph],
5.07 [d, 1H, C(1)-H], 6.98–7.66 [m, 22H, H–arom phenyl,
C(40)-H and C(50)-H], 9.53 (slarge, 1H, NH], J1,2=2.7,
J2,3=7.6, J3,4a=3.5, J3,4b=4.7, J3,OH=6.3, J4a,4b=10.1.
Procedure C. To a stirred solution of crude 22
(477 mg) in anhydrous THF (10 mL) under argon was
added dropwise a 1 M solution of TBAF in THF
(1.5 mL, 2 equiv). The reaction mixture was stirred
overnight at rt. Workup and chromatography gave 23
(196 mg, 84%) as a colourless solid. Mpdec=126–127 ꢁC
20
1
(toluene). ½aŠD =+60 (c 2, CHCl3). H NMR (CDCl3,
250 MHz): d=2.97 [slarge, 1H, OH], 3.90 [dd, 1H, C(8)-
Hb], 3.97 [dd, 1H, C(8)-Ha], 4.51 [dt, 1H, C(5)-H], 4.60
and 4.73 [AB, 2H, J=11.7, OCH2Ph], 4.72 [dd, 1H, C(6)-
H], 4.83 and 5.07 [AB, 2H, J=11.6, OCH2Ph], 4.91 [d,
1H, C(7)-H], 6.97 [d, 1H, C(2)-H or C(3)-H], 7.14 [d, 1H,
C(3)-H or C(2)-H], 7.27–7.46 [m, 10H, H–arom phenyl];
J2,3=0.9, JHa,Hb=12.2, JHa,5=4.3, JHb,5=4.3, J5,6=6.8,
J6,7=3.4. 13C NMR (CDCl3, 62.9 MHz]: d=59.3 [C(5)],
61.7 [CH2OH], 71.4 [OCH2Ph], 72.8 [OCH2Ph], 76.8
[C(7)], 87.4 [C(6)], 114.1 [C(3)], 127.9–128.7 [C–arom
phenyl], 133.6 [C(2)], 136.8 and 137.6 [Cs phenyl], 150.4
[C(7a)]. C21H22N2O3 (350.4): calcd C 71.98, H 6.33, N
7.99; found C 72.1, H 6.2, N 7.9.
1,2-di-O-Benzyl-4-O-TBDPS D-arabino derivative ent-
19. Procedure A, starting from a solution of ent-18,
mp=112–113 ꢁC7 (379 mg, 1.03 mmol), imidazole
(100 mg), DMAP (4.7 mg), Et3N (194 mL) and
TBDPSCl (325 mL, 1.21 mmol) in CH2Cl2 (12 mL).
After 48 h, workup and chromatographic purification
led to ent-19 (647 mg, 75%) as a colourless foam. H
NMR (CDCl3, 250 MHz) spectrum identical to, and
superimposable on, that of 19.
1
1,2-di-O-Benzyl-4-O-TBDPS L-ribo derivative 21. Pro-
cedure A as above, starting from 20, mp=111–112 ꢁC7
(525 mg, 1.42 mmol), imidazole (150 mg), DMAP
(8 mg), Et3N (270 mL) and TBDPSCl (470 mL, 1.75 mol)
in CH2Cl2 (15 mL). After 48 h, workup and chromato-
graphic purification led to 21 (736 mg, 85%) as a col-
6,7-di-O-Benzyl-8-OTBDPS L-xylo derivative ent-22
and 6,7-di-O-benzyl-L-xylo derivative ent-23. Procedure
B as above, starting from a solution of ent-19 (469 mg,
0.77 mmol), pyridine (245 mL, 3.09 mmol), in CH2Cl2
(12 mL) at ꢀ10 ꢁC to which Tf2O (390 mL, 2.32 mmol)
was added. After 12 h at rt workup and purification by
chromatography (AcOEt/cyclohexane 1:1) as above led
to ent-22 (330 mg) which was used as such in the next step.
1H NMR (CDCl3, 250 MHz) spectrum of ent-22 identical
to, and superimposable on, that of compound 22.
1
ourless foam. H NMR (CDCl3, 250 MHz): d=1.05 [s,
9H, (CH3)3C], 3.40 [slarge, OH], 3.42 [ddd, 1H, C(3)-H],
3.71 [dd, 1H, C(4)-Hb], 3.78 [dd, 1H, C(4)-Ha], 4.08 [dd,
1H, C(2)-H], 4.46 and 4.55 [AB, 2H, J=11.8, OCH2Ph],
4.56 and 4.88 [AB, 2H, J=10.9, OCH2Ph], 5.11 [d, 1H,
C(1)-H], 6.93 [slarge, 1H, C(40)-H or C(50)-H], 7.02 [slarge
,
1H, C(50)-H or C(40)-H], 7.13–7.45 and 7.62–7.65 [m,
20H, H–arom phenyl], 9.63 [slarge, NH], J1,2=2.3,
J2,3=8.7, J3, 4a=3.5, J3,4b=5.1, J4a,4b=10.4.
1,2-di-O-Benzyl-4-O-TBDPS D-ribo derivative ent-21.
Procedure
Procedure C. A solution of compound ent-22 (330 mg)
in THF (10 mL) was desilylated as above with a 1 M
A
as above, starting from ent-20,

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3567
solution of TBAF in THF (1.7 mL, 1.7 mmol). Workup
and purification by chromatography led to ent-23
(95 mg, 49%; overall yield from ent-19: 29%) as a col-
ourless solid. Mp=123 ꢁC. ½aŠD =ꢀ58 (c 2, CHCl3). H
NMR (CDCl3, 250 MHz) and 13C NMR (CDCl3,
62.9 MHz) spectra of ent-23 identical to, and super-
imposable on, those of 23. HR-MS [M+H]+ ion
351.1706 (C21H23N2O3, calcd 351.1709).
(1.32 mL, 2 equiv). The reaction mixture was stirred for
2 h at rt. Workup as above followed by chromatography
led to ent-25 as a colourless solid (91 mg, 39% from ent-
21). Mp=99.5 ꢁC (toluene). ½aŠD20=+135 (c 2; CHCl3).
1H NMR (CDCl3, 250 MHz) and 13C NMR (CDCl3,
62.9 MHz) spectra identical to, and superimposable on,
those of 25. HR-MS [M+H]+ ion 351.1708
(C21H23N2O3, calcd 351.1709).
20
1
6,7-di-O-Benzyl-8-OTBDPS D-lyxo derivative 24 and
6,7-di-O-benzyl D-lyxo derivative 25. Procedure B as
above starting from a solution of 21 (730 mg, 1.2 mmol),
pyridine (380 mL), in CH2Cl2 (15 mL) at ꢀ10ꢁ C to
which Tf2O (610 mL, 36 mmol) was added. After 12 h,
workup and purification by chromatography led to 24
(550 mg, 75%) which was used as such in the next step.
1H NMR (CDCl3, 250 MHz): d=1.07 [s, 9H, (CH3)3C],
4.01 [dd, 1H, C(4)-Hb], 4.13 [dd, 1H, C(4)-Ha], 4.31 [dd,
1H, C(6)-H], 4.39 and 4.58 [AB, 2H, J=12.2, OCH2Ph],
4.43 [ddd, 1H, C(5)-H], 4.57 [d, 1H, C(7)-H], 4.64 and
4.74 [AB, 2H, J=12.1, OCH2Ph], 7.13–7.17, 7.25–7.49
and 7.62–7.67 [m, 22H, H–arom phenyl and imidazole];
JHa,Hb=10.9, JHa,5=3.5, JHb,5=10.1, J5,6=7.1,
J6,7=5.2.
D-xylo-Imidazolo-pyrrolidinose 8. Procedure D, starting
from 23 (190 mg, 0.54 mmol) in MeOH (5 mL) under H2
pressure (30 bar) in the presence of 10% Pd/C contain-
ing 50% H2O (380 mg). After 3 days workup and chro-
matography led to 8 as a hygroscopic colourless powder
after lyophilisation (74 mg, 80%). Mpdec=173–174 ꢁC.
½aŠ20=+51 (c 1, MeOH). CD (H2O): recording starts at
D
180.0 (ꢀ9.80), 182.0 (ꢀ11.50), 195.0 (0.00), 206.0 (+
4.29), tailing out at 240.0 (0.0). 1H NMR (CD3OD,
400 MHz): d=3.85 [dd, 1H, C(8)-Hb], 3.94 [dd, 1H,
C(8)-Ha], 4.45 [td, 1H, C(5)-H], 4.61 [dd, 1H, C(6)-H],
4.82 [d, 1H, C(7)-H], 7.02 [d, 1H, C(2)-H], 7.12 [d, 1H,
C(3)-H], JHa,Hb=11.8, JHa,5=3.6, JHb,5=6.2, J5,6=6.2,
J6,7=4.0, J2,3=1.3. 13C NMR (CD3OD, 100.6 MHz): d
=61.6 [CH2OH], 62.4 [C(5)], 73.9 [C(7)], 82.9 [C(6)],
116.3 [C(3)], 132.5 [C(2)], 153.5 [C(7a)]. HR-MS:
[M+H]+ ion 171.0771 (C7H11N2O3, calcd 171.0770).
Procedure C. A solution of 24 (550 mg, 10.9 mmol) in
THF (5 mL) was desilylated as above with a 1 M solu-
tion of TBAF in THF (2 mL, 2.0 mmol). After 2 h,
standard workup and purification by chromatography
led to 25 (268 mg, 63% overall yield from 21) as a col-
ourless solid. Mpdec=104 ꢁC (toluene). ½aŠ2D0=ꢀ140 (c 2;
L-xylo-Imidazolo-pyrrolidinose ent-8. Procedure D,
starting from a solution of ent-23 (95 mg, 0.27 mmol) in
MeOH (1.5 mL) and AcOH (1.5 mL) under H2 pressure
(1.5 bar) in the presence of 20% Pd(OH)2/C (95 mg).
After 1 day workup and chromatography led to ent-8
after lyophilisation (36 mg, 78%) as a colourless pow-
der. Mpdec=163 ꢁC (MeOH); one of the monocrystals
was used for X-ray diffraction analysis (Fig. 2).
1
CHCl3). H NMR (CDCl3, 250 MHz): d=3.02 [dd, 1H,
OH], 3.92 [ddd, 1H, C(8)-Hb], 4.02 [dt, 1H, C(8)-Ha],
4.48 [ddd, 1H, C(5)-H] 4.52 and 4.79 [AB, 2H, J=11.5,
OCH2Ph], 4.55 [dd, 1H, C(6)-H], 4.73 [d, 1H, C(7)-H],
4.76 and 4.92 [AB, 2H, J=11.7, OCH2Ph], 7.02 [d, 1H,
C(3)-H], 7.19 [d, 1H, C(2)-H], 7.27–7.44 [m, 10H,
H–arom phenyl], J2,3=0.8, JHa,OH=3.7, JHb,OH=8.8,
½aŠ20=ꢀ55 (c 1, MeOH). CD (H2O): recording starts at
D
180.0 (+9.30), 182.0 (+11.37), 196.5 (0.00), 206.5
1
(ꢀ4.83), tailing out at 245.0 (0.0). H NMR (CD3OD,
250 MHz): d=3.84 [dd, 1H, C(8)-Ha], 3.93 [dd, 1H,
C(8)-Hb], 4.44 [td, 1H, C(5)-H], 4.61 [dd, 1H, C(6)-H],
4.82 [d, 1H, C(7)-H], 7.02 [d, 1H, C(2)-H], 7.12 [d, 1H,
C(3)-H], JHa,Hb=11.8, JHa,5=3.6, JHb,5=6.0, J6,7=4.0,
J5,6=6.3, J2,3=1.1. 13C NMR [CD3OD, 62.9 MHz): d
=61.6 [C(8)], 62.3 [C(5)], 73.9 [C(7)], 82.9 [C(6)], 116.2
[C(3)], 132.5 [C(2)], 153.50 [C(7a)]. These two NMR
spectra match those of compound 8. HR-MS: [M+H]+
ion 171.0771 (C7H11N2O3, calcd 171.0770).
JHa,Hb=12.0,
JHa,5=2.9,
JHb,5=4.1,
J5,6=7.3,
J6,7=5.2. 13C NMR (CDCl3, 62.9 MHz): d=59.9 [C(5)],
61.1 [C(8)], 68.0 [C(7)], 70.5 [OCH2Ph], 71.8 [OCH2Ph],
79.3 [C(6)], 115.0 [C(3)], 127.9-128.6 [C–arom phenyl],
133.1 [C(2)], 136.8 and 136.9 [Cs phenyl], 149.5 [C(7a)].
C21H22N2O3 (350.4): calcd C 71.98, H 6.33, N 7.99;
found C 71.9, H 6.2, N 7.9.
6,7-di-O-Benzyl-8-OTBDPS L-lyxo derivative ent-24
and 6,7-di-O-benzyl-L-lyxo derivative ent-25. Procedure
B. To a stirred solution of ent-21 (400 mg, 0.66 mmol),
and anhydrous pyridine (208 mL, 2.6 mmol) in CH2Cl2
(10 mL) at ꢀ10 ꢁC under argon was added dropwise
Tf2O (350 mL, 2.0 mmol). The reaction mixture was
stirred for 30 min at ꢀ10 ꢁC, then at rt. Workup led to
crude compound ent-24 (428 mg) which was used as
such for the next step. 1H NMR (CDCl3, 250 MHz)
spectrum of ent-24 identical to, and superimposable on
that of 24.
D-lyxo-Imidazolo-pyrrolidinose 9. Procedure D, starting
from 25 (250 mg, 0.71 mmol) in EtOH (10 mL) under H2
pressure (30 bar) in the presence of 10% Pd/C. After 5
days, standard workup, chromatography and lyophili-
sation led to 9 (90 mg, 74%) as a colourless solid. Mpdec
167–168 ꢁC. ½aŠD20=+36 (c 1, MeOH). CD (H2O):
recording starts at 180.0 (+5.89), 182.5 (+7.92), 193.5
(0.00), 198.0 (ꢀ1.85), 203.0 (0.00), 210.0 (+2.27), 224.0
(0.00), 230.0 (ꢀ0.30), tailing out at 245.0 (0.0). 1H NMR
(CD3OD, 400 MHz): d=3.81 [dd, 1H, C(8)-Hb], 3.93
[dd, 1H, C(8)-Ha], 4.40 [ddd, 1H, C(5)-H], 4.73 [dd, 1H,
C(6)], 4.76 [d, 1H, C(7)-H], 7.02 [d, 1H, C(2)-H], 7.12 [d,
1H, C(3)-H], JHa,Hb=11.7, JHa,5=3.4, JHb,5=5.6,
J5,6=6.3, 6,7=5.7, J2,3=1.3. 13C NMR (CD3OD,
100.6 MHz): d=61.2 [CH2OH], 62.8 [C(5)], 66.5 [C(7)],
Procedure C. To a stirred solution of crude ent-24
(428 mg) in anhydrous THF (11 mL) under argon was
added dropwise a 1 M solution of TBAF in THF

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Theophile Tschamber et al. / Bioorg. Med. Chem. 11 (2003) 3559–3568
3568
75.2 [C(6)], 116.4 [C(3)], 132.4 [C(2)], 153.7 [C(7a)]. HR-
MS: [M+H]+ ion 171.0771 (C7H11N2O3, calcd
171.0770).
Acknowledgements
We wish to thankthe Fondation pour l’Ecole de Chimie
de Mulhouse for a PhD grant to Francois Gessier.
Furthermore, the continuous support of the Centre
National de la Recherche Scientifique (UMR-7015) is
gratefully acknowledged.
L-lyxo-Imidazolo-pyrrolidinose ent-9. Procedure D,
starting from ent-25 (91 mg, 0.26 mmol) in MeOH
(1.5 mL) and AcOH (1.5 mL) under H2 atmosphere (ca.
1.2 bar) in the presence of Pd(OH)2/C (90 mg). After
20 h, workup and chromatography led to ent-9 after
lyophilisation (25 mg, 57%) as a colourless solid.
Mpdec=136 ꢁC. ½aŠD20=ꢀ35 (c 1, MeOH). CD (H2O):
recording starts at 180.0 (ꢀ5.57), 182.0 (ꢀ7.03), 193.5
(0.00), 198.0 (+1.23), 204.0 (0.00), 210.0 (ꢀ2.19), 227.0
References and Notes
1. Sinnott, M. L. Chem. Rev. 1990, 90, 1171.
2. Zechel, D. L.; Withers, S. G. Acc. Chem. Res. 2000, 33, 11.
3. Aoyagi, T.; Suda, H.; Uotani, K.; Kojima, F.; Aoyama, T.;
Horiguchi, K.; Hamada, M.; Takeuchi, T. J. Antibiot. 1992,
45, 1404.
4. Aoyama, T.; Naganawa, H.; Suda, H.; Uotani, K.; Aoyagi,
T.; Takeuchi, T. J. Antibiot. 1992, 45, 1557.
5. Tatsuta, K.; Miura, S.; Ohta, S.; Gunji, H. Tetrahedron
Lett. 1995, 36, 1085.
6. Tatsuta, K.; Miura, S.; Ohta, S.; Gunji, H. J. Antibiot.
1995, 48, 286.
7. Gessier, F.; Tschamber, T.; Tarnus, C.; Neuburger, M.;
Huber, W.; Streith, J. Eur. J. Org. Chem. 2001, 4111.
8. Heightman, T. D.; Vasella, A. T. Angew. Chem., Int. Ed.
1999, 38, 750.
1
(0.00), 231.5 (+0.11), tailing out at 243.0 (ꢀ0.10). H
NMR (CD3OD, 250 MHz): d=3.81 [dd, 1H, C(8)-Hb],
3.93 [dd, 1H, C(8)-Ha], 4.40 [ddd, 1H, C(5)-H], 4.70 [dd,
1H, C(6)-H], 4.76 [d, 1H, C(7)-H], 7.02 [d, 1H, C(2)-H],
7.12 [d, 1H, C(3)-H], JHa,Hb=11.7, JHa,5=3.4, JHb,5=5.6,
J5,6=5.7, J6,7=5.6, J2,3=1.1. 13C NMR (CD3OD,
62.9 MHz): d=61.1 [CH2OH], 62.7 [C(5)], 66.4 [C(7)],
75.1 [C(6)], 116.4 [C(3)], 132.3 [C(2)], 153.6 [C(7a)]. These
two NMR spectra match those of compound 9. HR-MS:
[M+H]+ ion 171.0766 (C7H11N2O3, calcd 171.0770).
X-ray diffraction analysis of ent-8
9. Lillelund, V. H.; Jensen, H. H.; Liang, X.; Bols, M. Chem.
Rev. 2002, 102, 515.
Single crystals of ent-8, suitable for X-ray crystal-
lography, were grown by crystallization from methanol.
Data were collected at 293 K on a Bruker-Nonius Kap-
paCCD area detector using Mo-Ka radiation
(l=0.71073). The compound with the chemical formula
of C7H10N2O3 crystallized in the orthorhombic space
group P212121. The dimensions of the unit-cell are:
a=4.8564(6), b=12.661(3), c=12.840(4), a=b=g=90ꢁ.
The usual corrections were applied. The structure was
solved using the program SIR 92.17 Anisotropic refine-
ment on all non-hydrogen atoms was carried out using
the program CRYSTALS.18 Scattering factors were
taken from the International Tables Vol. IV table 2.2B.
The plots were created using ORTEP-3 for Windows.19
10. Shibata, T.; Nakayama, O.; Tsurumi, Y.; Okuhara, M.;
Terano, H.; Kohsaka, M. J. Antibiot. 1988, 41, 296.
11. Tsujii, E.; Muroi, M.; Shiragami, N.; Takatsuki, A. Bio-
chem. Biophys. Res. Commun. 1996, 220, 459.
12. Merely for convenience, we use herein the ‘imidazolo-
pyrrolidinose’ short-hand notation. In actual fact, these sugar
derivatives are not aldopentoses, but rather analogues of the
corresponding lactones. See ref 8.
13. Tschamber, T.; Siendt, H.; Tarnus, C.; Deredas, D.;
Frankowski, A.; Kohler, J.; Streith, J. Eur. J. Org. Chem.
2002, 702.
14. Ichikawa, M.; Igarashi, Y.; Ichikawa, Y. Tetrahedron
Lett. 1995, 36, 1767.
15. La, Y.-T. J. Biol. Chem. 1967, 242, 5474.
16. Segel, I. H. Enzyme Kinetics; John Wiley & Sons: New
York, 1975; p 109.
17. Altomare, A.; Cascarano, G.; Giacovazzo, G.; Guagliardi,
A.; Burla, M. C.; Polidori, G.; Camalli, M. J. Appl. Cryst.
1994, 27, 435.
18. Watkin, D. J.; Prout, C. K.; Carruthers, J. R.; Betteridge,
P. W.; Cooper, R. I. CRYSTALS Issue 11; Chemical Crystal-
lography Laboratory: Oxford, UK, 2001.
CCDC 196227 contains the supplementary crystal-
lographic data for this paper. These data can be obtained
free of charge at http://www.ccdc.cam.ac.uk/conts/
retrieving.html (or from the Cambridge Crystal-
lographic Data Centre, 12, Union Road, Cambridge
CB2 1EZ, UK; fax: +44-1223-336033; e-mail: deposit
@ccdc.cam.ac.uk).
19. Farrugia, L. J. J. Appl. Cryst. 1997, 30, 565.