Synthesis and evaluation of calystegine B2 analogues as glycosidase inhibitors

Article abstract of DOI:10.1021/jo015639f

A practical synthesis of polyhydroxylated 6-oxa-nor-tropanes incorporating the essential structural features of calystegine B₂ from 5-deoxy-5-thioureido and 5-ureido-L-idofuranose precursors is presented. The methodology relies on the ability of pseudoamide-type nitrogen atoms (thiourea, urea, and carbamate) to undergo nucleophilic addition to the masked aldehyde group of the monosaccharide. The generated hemiaminal functionality may further undergo in situ intramolecular glycosidation to give the bicyclic aminoacetal compounds, the whole process being favored by the anomeric effect. A series of derivatives bearing different substituents at nitrogen has been prepared and screened against several glycosidases in comparison with xylonojirimycin-type piperidine analogues. Interestingly, strong and highly specific inhibition of bovine liver β-glucosidase was observed for 6-oxacalystegine B₂ analogues incorporating aromatic pseudoaglyconic groups. On the basis of these data, a 1-azasugar inhibition mode is proposed for this family of glycomimetics.

Full text of DOI:10.1021/jo015639f

7604
J . Org. Chem. 2001, 66, 7604-7614
Syn th esis a n d Eva lu a tion of Ca lystegin e B2 An a logu es a s
Glycosid a se In h ibitor s
M. Isabel Garc´ıa-Moreno,† J uan M. Benito,† Carmen Ortiz Mellet,*,† and
J ose´ M. Garc´ıa Ferna´ndez*,‡
Departamento de Quı´mica Orga´nica, Facultad de Qu´ımica, Universidad de Sevilla, E-41071 Sevilla,
Spain, and Instituto de Investigaciones Qu´ımicas, CSIC, Ame´rico Vespucio s/ n, Isla de la Cartuja,
E-41092 Sevilla, Spain
jogarcia@cica.es
Received March 20, 2001
A practical synthesis of polyhydroxylated 6-oxa-nor-tropanes incorporating the essential structural
features of calystegine B2 from 5-deoxy-5-thioureido and 5-ureido-L-idofuranose precursors is
presented. The methodology relies on the ability of pseudoamide-type nitrogen atoms (thiourea,
urea, and carbamate) to undergo nucleophilic addition to the masked aldehyde group of the
monosaccharide. The generated hemiaminal functionality may further undergo in situ intramo-
lecular glycosidation to give the bicyclic aminoacetal compounds, the whole process being favored
by the anomeric effect. A series of derivatives bearing different substituents at nitrogen has been
prepared and screened against several glycosidases in comparison with xylonojirimycin-type
piperidine analogues. Interestingly, strong and highly specific inhibition of bovine liver â-glucosidase
was observed for 6-oxacalystegine B2 analogues incorporating aromatic pseudoaglyconic groups.
On the basis of these data, a 1-azasugar inhibition mode is proposed for this family of glycomimetics.
In tr od u ction
peutic drugs for the treatment of viral infections,8 cancer,9
and metabolic disorders such as diabetes.10 On the other
hand, the occurrence of calystegines in the leaves, skins,
and sprouts of some edible vegetables such as potatoes,
egg plant, and sweet potato and their interaction with
mammalian liver glycosidases have raised concerns
regarding their safety in human diet.11
In contrast to the polyhydroxy pyrrolidine, piperidine,
pyrrolizidine, and indolizidine azasugar glycosidase in-
hibitors, which by now have been extensively investi-
gated, an understanding of the structural bases for
glycosidase inhibition by the calystegine group of alka-
loids has only been partially established. Even when they
may be viewed as hybrids of piperidine and pyrrolidine
Calystegines,1 bicyclic alkaloids that possess a nor-
tropane structure bearing hydroxyl groups varying in
position and stereochemistry,2 represent a recently dis-
covered group of plant secondary metabolites believed to
function as nutritional mediators in the plant rhizo-
sphere.3 Like other polyhydroxyalkaloids with structural
resemblance to sugars isolated from both plants and
microorganisms,4 iminosugars (“azasugars”),5 the calys-
tegines exhibit strong and specific glycosyl hydrolase
competitive inhibitory activity.6 They show, therefore,
considerable promise as probes for structure/function
studies of enzymatic mechanisms4,7 and as chemothera-
† Universidad de Sevilla. Phone: +34 954557150. Fax: +34
954624960. E-mail: mellet@cica.es.
(5) Although the term “azasugar” is widely used in the literature to
refer to glycomimetics where the endocyclic oxygen atom has been
replaced by nitrogen, the term is not strictly correct according to the
IUPAC-IUBM nomenclature recommendations for carbohydrates.
“Azahexose” would actually imply that a carbon atom has been
exchanged for a nitrogen. See: McNaught, A. D. Pure Appl. Chem.
1996, 68, 1919.
‡ Instituto de Investigaciones Qu´ımicas. Phone: +34 954489559.
Fax: + 34 954460565.
(1) The discoverers of these compounds named them “calystegins”.
However, in recent literature, the word is usually spelled “calystegines”
because the extra “e” brings the name into line with most other
alkaloids.
(2) For a specific review, see: Molyneux, R. J .; Nash, R. J .; Asano,
N. The Chemistry and Biological Activity of Calystegines and Related
Nortropane Alkaloids. In Alkaloids: Chemical and Biological Perspec-
tives; Pelletier, S. W., Ed.; Pergamon: Oxford, 1996; Vol. 11, p 303.
(3) (a) Tepfer, D.; Goldmann, A.; Pamboukdjian, N.; Maille, M.;
Lepingle, A.; Chevalier, D.; Denarie´, J .; Rosemberg, C. J . Bacteriol.
1988, 170, 1153. (b) Goldmann, A.; Message, B.; Tepfer, D.; Molyneux,
R. J .; Duclos, O.; Boyer, F.-D.; Pan, Y. T.; Elbein, A. D. J . Nat. Prod.
1996, 59, 1137.
(4) For recent reviews on alkaloid glycosidase inhibitors, including
the calystegines, see: (a) Asano, N.; Nash, R. J .; Molyneux, R. J .; Fleet,
G. W. J . Tetrahedron: Asymmetry 2000, 11, 1645. (b) Elbein, A. D.;
Molyneux, R. J . Alkaloid Glycosidase Inhibitors. In Comprehensive
Natural Products Chemistry; Barton, D., Nakanishi, K., Meth-Cohn,
O., Eds.; Elsevier: Oxford, 1999; Vol. 3, p 129. (c) Simmonds, M. S. J .;
Kite, G. C.; Porter, E. A. Taxonomic Distribution of Iminosugars in
Plants and Their Biological Activities. In Iminosugars as Glycosidase
Inhibitors; Stu¨tz, A., Ed.; Wiley-VCH: Weinheim, Germany, 1999; p
8. (d) Ossor, A.; Elbein, A. D. Glycoprotein Processing Inhibitors. In
Carbohydrates in Chemistry and Biology; Ernst, B., Hart, G. W., Sinay¨,
P., Eds.; Wiley-VCH: Weinheim, Germany, 2000; Part II, Vol. 3, p 513.
(6) (a) Asano, N.; Kato, A.; Miyauchi, M.; Kizu, H.; Tomimori, T.;
Matsui, K.; Nash, R. J .; Molyneux, R. J . Eur. J . Biochem. 1997, 248,
296. (b) Asano, N.; Kato, A.; Kizu, H.; Matsui, K.; Griffiths, R. C.; J ones,
M. G.; Watson, A. A.; Nash, R. J . Carbohydr. Res. 1997, 304, 173. (c)
Asano, N.; Kato, A.; Kizu, H.; Matsui, K.; Watson, A. A.; Nash, R. J .
Carbohydr. Res. 1996, 293, 195. (d) Asano, N.; Kato, A.; Yokoyama,
Y.; Miyauchi, M.; Yamamoto, M.; Kizu, H.; Matsui, K. Carbohydr. Res.
1996, 284, 169. (e) Asano, N.; Kato, A.; Oseki, K.; Kizu, H.; Matsui, K.
Eur. J . Biochem. 1995, 229, 369. (f) Asano, N.; Oseki, K.; Tomioka, E.;
Kizu, H.; Matsui, K. Carbohydr. Res. 1994, 259, 243. (g) Molyneux, R.
J .; Pan, Y. T.; Goldmann, A.; Tepfer, D. A.; Elbein, A. D. Arch. Biochem.
Biophys. 1993, 304, 81.
(7) For leading reviews, see: (a) Heightman, T. D.; Vasella, A. T.
Angew. Chem., Int. Ed. 1999, 38, 750. (b) Sears, P.; Wong, C.-H. Chem.
Commun. 1998, 1161. (c) Dwek, R. A. Chem. Rev. 1996, 96, 683-720.
(d) Kaushal, G. P.; Elbein, A. D. Methods Enzymol. 1994, 230, 316. (e)
Nishimura, Y. Glycosidase and Glycosyltransferase Inhibitors. In
Studies in Natural Products Chemistry; Atta-ur-Rahman, Ed.; Elsevi-
er: Amsterdam, 1992; Vol. 10, p 495. (f) Winchester, B.; Fleet, G. W.
J . Glycobiology 1992, 2, 199. (g) Legler, G. Adv. Carbohydr. Chem.
Biochem. 1990, 48, 319. (h) Sinnot, M. L. Chem. Rev. 1990, 90, 1171.
(i) Elbein, A. D. Annu. Rev. Biochem. 1987, 56, 497.
10.1021/jo015639f CCC: $20.00 © 2001 American Chemical Society
Published on Web 10/24/2001

Calystegine B2 Analogues
J . Org. Chem., Vol. 66, No. 23, 2001 7605
carbohydrate mimics, their enzyme specificities may be
drastically different. Thus, calystegine B2 (1), one of the
most powerful representatives, has topographical proper-
ties similar to those of the potent R-glycosidase inhibitors
1-deoxynojirimycin (2) and castanospermine (3). Not-
withstanding, compound 1 behaves instead as a potent
inhibitor of â-glucosidases, whereas little or no inhibition
is detected for R-glucosidases. A binding model has been
proposed that invokes charge interactions with the
catalytic carboxylic groups at the active site by analogy
with the presumed glycosyl oxocarbenium cation inter-
mediate of enzymatic glycoside cleavage (4).6e,12 However,
in light of recent mechanistic studies on the â-glucosidase
reaction,13 the remarkable â-anomeric selectivity would
rather suggest a correspondence with an anomeric car-
bocation species (5). Calystegine B2 might then be
regarded as a rigid analogue of the strong 1-azasugar
â-glucosidase inhibitor isofagomine (6),14 a stereochemical
mimic of D-glucose in which the anomeric carbon has been
replaced by nitrogen.15 Surprisingly, compound 1 is also
a potent inhibitor of several R-galactosidases, suggesting
that different recognition patterns may operate with
different enzymes.12 Confirmation of any inhibition model
has been prevented, however, by the absence of versatile
synthetic routes to calystegine analogues. The reported
approaches to the preparation of polyhydroxy-nor-tro-
panes are, generally, rather long,16,17 and no ring-modi-
fied analogues have been described so far.
the masked aldehyde group of a monosaccharide.18 The
conformational properties and reactivity of the generated
aminoketalic center were shown to be governed by the
generalized anomeric effect. Exclusively dispositions with
an axially oriented pseudoanomeric oxygen substituent
are allowed, probably due to a very efficient delocalization
interaction between the π-type lone-pair orbital of the
sp2-hybridized nitrogen atom in the ground state of
N-(thio)carbonyl functionalities and the σ*-antibonding
orbital of the contiguous C-O bond (Figure 1). We
assumed that the same orbital interactions should favor
intramolecular glycosidation of the hemiaminal function-
ality by suitably located OH groups provided that the
anomeric effect is fulfilled, giving access to bridged 1,3-
O,N-heterobicyclic systems.19 This principle was the
subject of a preliminary account20 and has now been
translated into a practical synthesis of 1-deoxy-6-oxaca-
lystegine B2 glucomimetics (Figure 2), a hitherto un-
known class of compounds, from (thio)carbamic (thiourea,
urea, carbamate) carbohydrate precursors. Preparation
of the key intermediates, the scope and limitations of the
methods, and the structure/glycosidase inhibitory selec-
tivities and potency relationships are discussed in com-
parison with data for monocyclic nojirimycin-type ana-
logues.21
We have previously reported a versatile synthesis of
reducing N-thiocarbonyl and N-carbonyl azasugar gly-
comimetics related to castanospermine by intramolecular
nucleophilic addition of (thio)carbamic-type groupings to
(8) (a) van den Broek, L. A. G. M. Azasugars: Chemistry and Their
Biological Activity as Potential Anti-HIV Drugs. In Carbohydrates in
Drug Design; Witczak, Z. J ., Nieforth, K. A., Eds.; Marcel Dekker: New
York, 1997; p 471. (b) Carlson, G. B.; Butters, T. D.; Dwek, R. A.; Platt,
F. M. J . Biol. Chem. 1993, 268, 570. (c) Taylor, D. L.; Sunkara, P. S.;
Liu, P. S.; Kang, M. S.; Bowlin, T. L.; Tyms, A. S. AIDS 1991, 5, 693.
(d) Sunkara, P. S.; Taylor, D. L.; Kang, M. S.; Bowlin, T. L.; Liu, P. S.;
Tyms, A. S.; Sjoerdsma, A. Lancet 1989, 1206. (e) Karpas, A.; Fleet,
G. W. J .; Dwek, R. A.; Petursson, S.; Namgoong, S. K.; Ramsden, N.
G.; J acob, G. S.; Rademacher, T. W. Proc. Natl. Acad. Sci. U.S.A. 1988,
85, 9229.
Resu lts
A retrosynthetic analysis revealed that the bicyclic
6-oxa-nor-tropane skeleton can be constructed by in-
tramolecular O-6-glycosylation of reducing azasugar-type
intermediates (Figure 2). Our synthetic strategy relies
on the ability of 5-deoxy-5-thioureido and 5-ureido al-
dohexofuranose derivatives (Figure 2, structure III) to
undergo spontaneous rearrangement to the correspond-
(9) For a review, see: Gross, P. E.; Baker, M. A.; Carver, J . P.;
Dennis, J . W. Clin. Cancer Res. 1995, 1, 935.
(10) (a) Witczak, Z. J . Carbohydrates as New and Old Targets for
Future Drug Design. In Carbohydrates in Drug Design; Witczak, Z.
J ., Ed.; Marcel Dekker: New York, 1997; p 1. (b) Balfour, J . A.;
McTavish, D. Drugs 1993, 46, 1025. (c) Robinson, K. M.; Begovic, M.
E.; Rhinehart, B. L.; Heineke, E. W.; Ducep, J .-B.; Kastner, P. R.;
Marshall, F. N.; Danzin, C. Diabetes 1991, 40, 825. (d) Anzeveno, P.
B.; Creemer, L. J .; Daniel, J . K.; King, C.-H.; Liu, P. S. J . Org. Chem.
1989, 54, 2539. (e) Platt, F. M.; Neises, G. R.; Reinkensmeier, G.;
Townsend, M. J .; Perry, V. H.; Proia, R. L.; Winchester, B.; Dwek, R.
A.; Butters, T. D. Science 1997, 276, 428.
(11) Asano, N.; Kato, A.; Matsui, K.; Watson, A. A.; Nash, R. J .;
Molyneux, R. J .; Hackett, L.; Topping, J .; Winchester, B. Glycobiology
1997, 7, 1085.
(12) Legler, G. Glycosidase Inhibition by Basic Sugar Analogs and
the Transition State of Enzymatic Glycoside Hydrolysis. In Iminosug-
ars as Glycosidase Inhibitors; Stu¨tz, A., Ed.; Wiley-VCH: Weinheim,
Germany, 1999; p 31.
(13) (a) Rye, C. S.; Withers, S. G. Curr. Opin. Chem. Biol. 2000, 4,
573. (b) Namchuck, M. N.; Withers, S. G. Biochemistry 1995, 34, 16194.
(c) Tull, D.; Withers, S. G. Biochemistry 1994, 33, 16194.
(14) (a) J espersen, T. M.; Dong, W.; Sierks, M. R.; Skrydstrup, T.;
Lundt, I.; Bols, M. Angew. Chem., Int. Ed. Engl. 1994, 33, 1778. (b)
J espersen, T. M.; Bols, M.; Sierks, M. R. Tetrahedron 1994, 50, 13449.
(c) Dong, W.; J espersen, T. M.; Bols, M.; Skrydstrup, T.; Sierks, M. R.
Biochemistry 1996, 35, 2788. (d) Ichikawa, Y.; Igarashi, Y.; Ichikawa,
M.; Suhara, Y. J . Am. Chem. Soc. 1998, 120, 3007.
(15) For recent reviews on 1-azasugars, see: (a) Lundt, I.; Madsen,
R. Isoiminosugars: Glycosidase Inhibitors with Nitrogen at the Ano-
meric Position. In Iminosugars as Glycosidase Inhibitors; Stu¨tz, A.,
Ed.; Wiley-VCH: Weinheim, Germany, 1999; p 112. (b) Bols, M. Acc.
Chem. Res. 1998, 31, 1.
(16) (a) Ducrot, P.-H.; Beauhaire, J .; Lallemand, J .-Y. Tetrahedron
Lett. 1990, 31, 969. (b) Boyer, F.-D.; Lallemand, J .-Y. Synlett 1992,
969. (c) Boyer, F.-D.; Ducrot, P.-H.; Henryon, V.; Soulie´, J .; Lallemand,
J .-Y. Synlett 1992, 357. (d) Duclos, O.; Mondange, M.; Dure´ault, A.;
Depezay, J . C. Tetrahedron Lett. 1992, 33, 8061. (e) Duclos, O.;
Dure´ault, A.; Depezay, J . C. Tetrahedron Lett. 1992, 33, 1059. (f) Boyer,
F.-D.; Lallemand, J .-Y. Tetrahedron 1994, 50, 10443. (g) Soulie´, J .;
Faitg, T.; Betzer, J .-F.; Lallemand, J .-Y. Tetrahedron 1996, 52, 15137.
(h) Faitg, T.; Soulie´, J .; Lallemand, J .-Y.; Ricard, L. Tetrahedron:
Asymmetry 1999, 10, 2165.
(17) For a notably short synthesis of calystegine B2, see: Boyer, F.-
D.; Hanna, I. Tetrahedron Lett. 2001, 42, 1275.
(18) (a) D´ıaz Pe´rez, V. M.; Garc´ıa-Moreno, M. I.; Ortiz Mellet, C.;
Fuentes, J .; D´ıaz Arribas, J . C.; Can˜ada, F. J .; Garc´ıa Ferna´ndez, J .
M. J . Org. Chem. 2000, 65, 136. (b) J ime´nez Blanco, J . L.; D´ıaz Pe´rez,
V. M.; Ortiz Mellet, C.; Fuentes, J .; Garc´ıa Ferna´ndez, J . M.; D´ıaz
Arribas, J . C.; Can˜ada, F. J . Chem. Commun. 1997, 1969.
(19) J ime´nez Blanco, J . L.; Ortiz Mellet, C.; Fuentes, J .; Garc´ıa
Ferna´ndez, J . M. Tetrahedron 1998, 54, 14123.
(20) Garc´ıa Ferna´ndez, J . M.; Ortiz Mellet, J . M.; Benito, J . M.;
Fuentes, J . Synlett 1998, 316.
(21) Garc´ıa-Moreno, M. I.; Ortiz Mellet, C.; Garc´ıa Ferna´ndez, J .
M. Tetrahedron: Asymmetry 1999, 10, 4271.

7606 J . Org. Chem., Vol. 66, No. 23, 2001
Garc´ıa-Moreno et al.
Sch em e 1a
F igu r e 1. Stereochemical outcome of the intramolecular
nucleophilic addition of (thio)carbamic-type nitrogen atoms to
carbonyl groups (X ) S, O).
F igu r e 2. Structures of (I) 6-oxacalystegine, (II) polyhydrox-
ypiperidine, and (III) aldohexofuranose.
ing polyhydroxylated N-(thio)carbamoyl piperidines (Fig-
ure 2, structure II) through the open-chain tautomeric
form, with simultaneous generation of the reactive ami-
noacetal group. A hydroxylation profile analogous to that
of calystegine B2 (1) in the final compounds (Figure 2,
structure I) implies an L-ido configuration of the (thio)-
urea precursors.
a
Reagents: (i) RNCS (R ) Fmoc, Me, Bn, Ph), pyridine, Et3N;
(ii) pyridine, Et3N; (iii) 20% piperidine in MeOH, CH2Cl2 (60%).
sponding isothiocyanate gave the carbodiimide adducts
18-20. The yield varied from 80-90% for isothiocyanates
bearing electron-withdrawing groups (i.e., phenyl and
glucopyranosyl) to 31% for the alkyl-type isothiocyanate
9. Acid-catalyzed nucleophilic addition of water to the
carbodiimide group afforded the 5-deoxy-5-ureido-L-id-
ofuranose derivatives 21-23 (Scheme 2).26
Conventional deacetylation and/or acid hydrolysis of
the acetal protecting group in 11-16 and 21-23 with
TFA-water led, initially, to R,â-anomeric mixtures of the
corresponding L-idofuranose derivatives, with the (thio)-
urea group probably being protonated, as seen from the
13C NMR and FABMS spectra of the crude reaction
mixtures. Upon neutralization with Amberlite IRA 68
(OH-) ion-exchange resin, the formed N-(thio)carbam-
oylpiperidine derivatives underwent in situ intramolecu-
lar glycosylation reaction involving OH-6 to give the
target trihydroxylated N-(thio)carbamoyl-6-oxa-nor-tro-
pane glucomimetics 24-32 (Scheme 3).27,28 A similar pH-
dependent equilibrium between hemiacetal and hemi-
Syn th esis a n d Str u ctu r e of 6-Oxa -n or -tr op a n es.
The initial synthetic objective of this research was the
preparation of N-thiocarbamoyl and N-carbamoyl-6-ox-
acalystegine B2 derivatives bearing different substituents
at the exocyclic N′-atom. Reaction of 5-amino-5-deoxy-
1,2-O-isopropylidene-â-L-idofuranose (7), available in a
multigram scale (five steps) from commercial D-glu-
curono-6,3-lactone,22 with 9-fluorenylmethoxycarbonyl
(Fmoc), methyl, benzyl, and phenyl isothiocyanates af-
forded the required thioureas 10-13 with total chemose-
lectivity. Analogously, nucleophilic addition of amine 7
to 2,3,4,6-tetra-O-acetyl-â-D-glucopyranosyl isothiocyan-
ate (8)23 and methyl 2,3,4-tri-O-acetyl-6-deoxy-6-isothio-
cyanato-R-D-glucopyranoside (9)24 yielded the (1 f 5) and
(6 f 5) thiourea-bridged pseudodisaccharides 15 and 16,
respectively (Scheme 1).25 The N-monosubstituted thio-
urea 14 was obtained from 10 by removal of the N′-Fmoc
protecting group with piperidine.
For the preparation of the urea analogues, a different
synthetic strategy that avoids the use of hazardous
isocyanate reagents was devised. Condensation of the
per-O-protected azide 17 with triphenylphosphine and
further in situ aza-Wittig-type reaction with the corre-
(26) For examples of aza-Wittig-type transformations in carbohy-
drate chemistry, see: (a) D´ıaz Pe´rez, V. M.; Ortiz Mellet, C.; Fuentes,
J .; Garc´ıa Ferna´ndez, J . M. Carbohydr. Res. 2000, 326, 161. (b) Garc´ıa
Ferna´ndez, J . M.; Ortiz Mellet, C.; D´ıaz Pe´rez, V.; Fuentes, J .; Kova´cs,
J .; Pinte´r, I. Carbohydr. Res. 1997, 304, 261. (c) Garc´ıa Ferna´ndez, J .
M.; Ortiz Mellet, C.; D´ıaz Pe´rez, V.; Fuentes, J .; Kova´cs, J .; Pinte´r, I.
Tetrahedron Lett. 1997, 38, 4161.
(22) Dax, K.; Gaigg, B.; Grassberger, V.; Ko¨lblinger, B.; Stu¨tz, A.
E. J . Carbohydr. Chem. 1990, 9, 479.
(23) Camarasa, M. J .; Ferna´ndez-Resa, P.; Garc´ıa-Lo´pez, M. T.; de
las Heras, F. G.; Me´ndez-Castrillo´n, P. P.; San Fe´lix, A. Synthesis 1984,
509.
(24) Garc´ıa Ferna´ndez, J . M.; Ortiz Mellet, C.; Fuentes, J . J . Org.
Chem. 1993, 58, 5192.
(25) For reviews on thiocarbonyl carbohydrate derivatives, see: (a)
Garc´ıa Ferna´ndez, J . M.; Ortiz Mellet, C. Adv. Carbohydr. Chem.
Biochem. 1999, 55, 35. (b) Garc´ıa Ferna´ndez, J . M.; Ortiz Mellet, C.
Sulfur Rep. 1996, 19, 61.
(27) The 13C NMR spectra of the crude reaction mixtures in D2O
showed signals at 102.1-101.8 and 96.1-95.9 ppm, with similar
intensities, for the C-1 resonances of the R- and â-idofuranose anomers,
respectively. See: Bock, K.; Pedersen, C. Adv. Carbohydr. Chem.
Biochem. 1983, 41, 27. Upon addition of 0.1 M NaOD to the solution
in the NMR tube until a neutral to slightly basic pH was achieved, an
instantaneous, virtually quantitative transformation into the final
bicyclic compound was observed. The pseudomolecular peaks in the
FABMS spectra of the crude reaction mixtures before neutralization
showed an 18 m/z unit increase compared to those of the final
compounds, in agreement with the proposed reaction pathway.

Calystegine B2 Analogues
Sch em e 2
J . Org. Chem., Vol. 66, No. 23, 2001 7607
Sch em e 3a
aminalforms has previouslybeen observed for polyhydroxy-
γ-oxoguanidines.29
a
Reagents: (i) Amberlite IRA 68 (OH-) (60-96%).
To compare both the reactivity and biological proper-
ties of pseudoamide vs amine-type 6-oxacalystegine
analogues, preparation of the N-unsubstituted 1-deoxy-
6-oxacalystegine B2 35 was of interest. Attempts to obtain
this compound directly from amine 7 or its N-Boc- or
N-Fmoc-protected derivatives by acid treatment failed,
probably due to further hydrolysis-dehydration reactions
of the transient aminoacetal derivative in acidic media.30
Instead, the benzyl carbamate 3331 was prepared that,
after acid hydrolysis of the isopropylidene group and
neutralization as described above, afforded the N-bezy-
loxycarbonyl-6-oxa-nor-tropane 34.32 Hydrogenolysis of
34 led to the target compound 35 in 60% yield (Scheme
4).
Sch em e 4a
The 1H NMR spectra of 24-32, 34, and 35 showed
vicinal coupling constants characteristic of trans-diaxial
a
Reagents: (i) benzylchloroformate, Na2CO3 (85%); (ii) 90%
TFA-H2O, Amberlite IRA 68 (OH-) (80%); (iii) H2, 10% Pd/C
(60%).
(28) No products arising from inter- or intramolecular addition of
sulfur to the carbonyl group of the monosaccharide were detected.
Although a rationalization of the sulfur vs nitrogen nucleophilicity in
thioureas is problematic, from the ensemble of results available in the
literature (cf. ref 24), it appears that nitrogen is generally involved in
nucleophilic additions to carbonyl groups, whereas nucleophilic dis-
placement reactions generally proceed through sulfur.
(29) J eong, J .-H.; Murray, B. W.; Takayama, S.; Wong, C.-H. J . Am.
Chem. Soc. 1996, 118, 4227.
(30) (a) Paulsen, H.; Todt, K. Chem. Ber. 1967, 100, 512. (b) Paulsen,
H.; Todt, K. Angew. Chem. 1965, 77, 589.
(31) Schaller, C.; Vogel, P.; J a¨ger, V. Carbohydr. Res. 1998, 314,
25.
(32) The 5-benzyloxycarbonylamino-5-deoxy-â-L-idofuranose struc-
ture has been previously assigned to the product of acid hydrolysis of
acetonide 33 (cf. ref 31). However, the NMR data (CD3OD) provided
by the authors are rather consistent with a piperidine derivative (low-
field-shifted δC-1 in agrement with the existence of an N-C-1 bond),
in a chair conformation (J 2,3 ) 8.0 Hz, J 3,4 ) 8.1 Hz), bearing an axial
anomeric substituent (J 1,2 ) 1.7 Hz). The isolated compound was most
likely a hydrated form of 34.
dispositions for the H-2, H-3, and H-4 protons, with the
bridgehead H-1 and H-5 protons in equatorial orienta-
tions,33 in agreement with the chair conformation for the
six-membered ring. In addition, long-range 4J H,H coupling
constants between protons in a W-arrangement, e.g., H-4/
H-6exo and H-1/H-5, were also observed, as expected for
the rigid bicyclo[3.2.1]octane skeleton. The low-field 13C
chemical shift of C-6 confirmed its involvement in the
oxazolidine ring closure.33
(33) For clarity of presentation, the authors chose not to use the
numbers resulting from the heterocyclic compounds (see Experimental
Section) in the notation of atoms for NMR data. Instead, the notation
is kept consistent with the parent carbohydrate compounds. See Figure
2 for atom notation equivalency.

7608 J . Org. Chem., Vol. 66, No. 23, 2001
Garc´ıa-Moreno et al.
Sch em e 5a
Syn th esis a n d Str u ctu r e of N-Th ioca r ba m oyl a n d
N-Ca r ba m oyl P ip er id in es. To determine the influence
of the C-1-C-5 two-atom bridge on the inhibitory proper-
ties of this family of glycomimetics, the preparation and
biological evaluation of a series of monocyclic piperidine
analogues seemed intriguing. Attempts to prepare these
compounds from 1,2-O-isopropylidene-R-D-xylofuranose
precursors34 following the above synthetic strategy failed,
however, due to extensive decomposition of the products
upon acid treatment. An alternative route was developed
on the basis of the use of peracetylated xylofuranose
templates. Replacement of the acetonide group by acetyl
in the known azide 3635 afforded the triacetate 37 as an
inseparable mixture of the R- and â-anomers. Condensa-
tion of 37 with triphenylphosphine and subsequent aza-
Wittig-type reaction with either carbon disulfide or
methyl, phenyl, or 2,3,4,6-tetra-O-acetyl-â-D-glucopyra-
nosyl isothiocyanate led to the isothiocyanate 38 or
carbodiimide derivatives 42-44, respectively. Nucleo-
philic addition of methylamine, aniline, and 2,3,4,6-tetra-
O-acetyl-â-D-glucopyranosylamine36 to 38 yielded the
thiourea adducts 39-41, respectively, whereas acid-
catalyzed addition of water to the heteroallene group of
42-44 afforded the corresponding oxocompounds 45-47
(Scheme 5).
Deacetylation of 39-41 and 45-47 in methanol under
standard NaOMe-catalyzed conditions proceeded with
concomitant rearrangement to the target polyhydroxypi-
peridine tautomers. In the case of the thiourea deriva-
tives, subsequent fast glycosidation at the aminoacetal
center occurred, even using short reaction times and low
temperatures, leading to the R-methyl glycosides 48-
50.37 The later reaction was much slower for the urea
analogues. The use of short reaction times allowed the
reducing xylonojirimycin-type azaheterocycles 51-53 to
be isolated in 60-70% yields (Scheme 5).
Both the N-thiocarbamoyl and N-carbamoyl com-
pounds 48-50 and 51-53, respectively, existed in D2O
solution as single diastereomers. The high-field shift of
the C-1 resonance33 confirmed the aminoacetal and
a
Reagents: (i) 50% TFA-H2O, 5 h; (ii) 1:1 Ac2O-pyridine, 12
h (70% overall); (iii) TPP, CS2, dioxane (95%); (iv) RNH2 (70-95%);
(v) RNCS, TPP, toluene, 24 h (45-87%); (vi) 2:1 acetone-H2O,
1% TFA (60-95%); (vii) NaOMe, MeOH, then Amberlite IR-120
(H+) (54-88%).
3
hemiaminal structure, whereas the vicinal J H,H values
around the piperidine ring unambiguously pointed to a
conformation close to a chair and to the R configuration
for the new stereocenter, with the pseudoanomeric meth-
oxy or hydroxy group in the axial position, fitting the
anomeric effect.
Biologica l Activity. The inhibitory activities of the
oxacalystegine B2 derivatives 24-32, 34, and 35 and of
the monocyclic piperidine analogues 48-53 for R-glu-
cosidase (yeast), â-glucosidase (almonds), â-glucosidase
(bovine liver, cytosolic), and R-galactosidase (green coffee
beans) are summarized in Table 1. None of these com-
pounds inhibited R-glucosidase, in agreement with the
linkage specificity of the parent calystegine alkaloids. The
bicyclic N-thiocarbamoyl nor-tropane derivatives 25 and
26 acted as competitive inhibitors of both the almond and
the cytosolic â-glucosidases, the inhibition potency being
about 2-3 orders of magnitude higher for the mam-
malian enzyme. It may be concluded that these types of
compounds can discriminate not only between R- and
â-glucosidases but also between â-glucosidases of a
different origin. A strong influence of the nature of the
N′-substituent on the inhibition constant was observed,
the potency as an inhibitor of â-glucosidase being about
100-fold higher for compounds bearing aromatic substit-
uents than for compounds with methyl, glucosyl, or
hydrogen substituents. The inhibition potency also in-
creased by a factor of about 10 on going from N-carbamoyl
(30) to N-thiocarbamoyl derivatives (25 and 26). Actually,
the N-(N′-phenyl)thiocarbamoyl derivative 26 inhibits the
mammalian cytosolic â-glucosidase (Ki ) 2.5 µM) 18-fold
more strongly than the natural compound calystegine B2
1 (Ki ) 45 µM). When considering the almond â-glucosi-
dase, we found that the corresponding Ki values (970 and
1.5 µM for 26 and 1, respectively) indicated a reverse
specificity. Conversely, the unsubstituted derivative 35,
having an amine-type endocyclic nitrogen, exhibited a
behavior toward this particular pair of â-glucosidases
parallel to that of calystegine B2. The 10-fold increment
in the corresponding inhibition constant values of 35 (85
and 606 µM for the almond and bovine liver enzymes,
(34) Garc´ıa-Moreno, M. I.; Benito, J . M.; Ortiz Mellet, C.; Garc´ıa
Ferna´ndez, J . M. Tetrahedron: Asymmetry 2000, 11, 1331.
(35) Szarek, W. A.; J ones, J . K. N. Can. J . Chem. 1965, 43, 2345.
(36) Babiano Caballero, R.; Fuentes Mota, J .; Ga´lbis Pe´rez, J . A.
Carbohydr. Res. 1986, 154, 280.
(37) Deacetylation of 40 was also carried out using NaCD3 in
CD3OD. The corresponding NMR spectra of the mixture evidenced the
formation of the corresponding deuterated methyl glycoside under
these conditions, discarding an alternative acid-catalyzed mechanism
during neutralization with the resin.

Calystegine B2 Analogues
J . Org. Chem., Vol. 66, No. 23, 2001 7609
Ta ble 1. Com p a r ison of In h ibitor y Activities (Ki, µM) for Bicyclic (24-32, 34, a n d 35) a n d Mon ocyclic Glycom im etics
Rela ted to Ca lystegin e B2 (48-53)
enzyme
enzyme
R-glucosidase â-glucosidase â-glucosidase R-galactosidase
R-glucosidase â-glucosidase â-glucosidase R-galactosidase
compd
(yeast)
(almonds)
(bovine liver)
(green coffee)
compd
(yeast)
(almonds)
(bovine liver)
(green coffee)
24
25
26
27
28
29
30
31
32
n.i.a
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
300
5.3
2.5
n.i.
148
325
30
n.i.
1640
n.i.
163
137
170
n.i.
n.i.
172
160
175
34
35
48
49
50
51
52
53
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
85
70
17800
191
n.i.
400
n.i.
n.i.
550
n.i.
1500
970
n.i.
606
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
n.i.
1500
n.i.
n.i.
1500
n.i.
n.i.
118
1500
n.i.
n.i.
a
No inhibition (n.i.) detected.
is probably the reason that the hemiaminal compounds
51-53 could be isolated in the D-xylose-derived N-
carbamoyl piperidine series, whereas the thioxo coun-
terparts underwent in situ glycosidation by methanol (f
48-50).39 The anomeric effect stabilization also explains
the high stability of all the N-(thio)carbamoyl azasugars
reported here as well as their configurational and con-
formational integrity, which is notably different than that
found for the amine derivative 35 due to the lability of
the N/O acetal function.40 Thus, whereas 26-32 and 48-
53 were found to be stable for weeks as solids or as
aqueous solutions in a refrigerator, compound 35 under-
went extensive decomposition after 1 day.
F igu r e 3. Anomeric-effect-driven intramolecular glycosylation
in L-ido-configurated polyhydroxypiperidine intermediates.
respectively), in comparison with the corresponding
values for the parent natural alkaloid, probably reflects
the contribution of the bridgehead hydroxy group of
calystegines to the binding energy. In stark contrast, the
monocyclic counterparts did not significantly affect any
of the glucosidases considered in this study.
Azasugars such as 1-deoxynojirimycin (2) are thought
to be good but rather nonspecific inhibitors of glycosi-
dases because they mimic the glycosyl oxocarbenium
cation (e.g., 4), a proposed intermediate in the mechanism
of action of both R- and â-glycosidases, upon protonation.
In contrast, 1-azasugars such as isofagomine (6) would
be good mimics of the glycosyl carbocation (e.g., 5), the
proposed first transition state in the mechanism of
â-glycosidases.14d In N-(thio)carbonyl compounds, the
basicity at the nitrogen atom is drastically decreased (by
14 pK units) compared to that in the corresponding amine
while keeping a positive charge density that results from
delocalization of the lone electron pair into the (thio)-
carbonyl group.38,41 This scenario is probably closer to
that encountered in the transition state of enzymatic
glycoside hydrolysis at the anomeric region.12,42 One could
expect that if glycomimetics of the calystegine family
behave as inhibitors of the 1-azasugar type, substituents
at the heterocyclic nitrogen would project into the aglycon
The strength of inhibition of R-galactosidase by this
family of compounds was, in general, much less influ-
enced by the nature of the N′-substituent. Values in the
137-191 µM range, much higher than the reported data
for calystegine B2 (0.86 µM), were measured for deriva-
tives bearing lipophilic or hydrophilic substituents in both
the thiocarbamoyl and carbamoyl series, as well as for
the unsubstituted compound 35.
Discu ssion
We have designed and synthesized a new class of
glycosidase inhibitors, the polyhydroxy 6-oxa-nor-tro-
panes. The synthetic scheme employs N-(thio)carbamic
monosaccharide precursors and involves a tandem fura-
nose f piperidine rearrangement-intramolecular gly-
cosylation process. The L-ido configuration considered in
this study leads to compounds that incorporate the
essential structural features of the azasugar alkaloid
calystegine B2 (1), i.e., an 8-azabicyclo[3,2,1]octane sys-
tem and an identical orientational pattern for the hy-
droxy groups at C-2, C-3, and C-4, with the hemiaminal
functionality characteristic of the natural compound
being replaced by an intramolecular aminoacetal group-
ing. To the best of our knowledge, these are the first
examples of ring-modified calystegine analogues.
The propensity of the reducing N-(thio)carbamoylpip-
eridine intermediate to undergo glycosidation reactions
is in agreement with formation of a transient azacarbe-
nium cation and an efficient stabilization of the transition
state leading to aminoacetal formation by the incipient
anomeric effect, the whole pathway being favored by the
π-symmetry of the lone-pair orbital of the N-atom in
pseudoamide functional groups (Figure 3). Because the
partial double-bond character of the N-C(X)N bond
decreases on going from thiourea to urea, a parallel
abatement of the contribution of orbital interactions to
the generalized anomeric effect was expected.25,38 This
(38) For reviews on the comparative chemical and electronic proper-
ties of N-(thio)carbonyl compounds, see: (a) Molina, M. T.; Ya´n˜ez, M.;
Mo´, O.; Notario, R.; Abboud, J .-L. M. The Thiocarbonyl Group. In
Supplement A3, The Chemistry of Double-Bonded Functional Groups;
Patai, S., Ed.; J ohn Wiley & Sons: Chichester, England, 1997; Part 2,
p 1355. (b) Duus, F. Thiocarbonyl Compounds. In Comprehensive
Organic Chemistry; Barton, D., Ollis, W. D., Eds.; Pergamon Press:
London, 1979; Vol. 3, p 373.
(39) The formation of the methyl glycosides 48-50 under basic
conditions is noteworthy. Glycosylation reactions generally need acid
catalysis. The observed reactivity can be rationalized assuming that
the transient hemiaminal precursor is in equilibrium with the corre-
sponding ∆1-piperidinium cation, which undergo electrophilic adition
of methylate. A similar mechanism has been put forward by other
authors to explain the formation of glycosides from heterocyclic
hemiaminals under neutral conditions. See: (a) Izquierdo, I.; Plaza,
M. T.; Robles, R.; Mota, A. J . Eur. J . Org. Chem. 2000, 2071. (b)
Dom´ınguez, M.-J .; Garc´ıa Lo´pez, M.-T.; Herranz, R.; Mart´ın Mart´ınez,
M.; Goza´lez Mun˜iz, R. J . Chem. Soc., Perkin Trans. 1 1995, 2839. (c)
Dom´ınguez, M.-J .; Garc´ıa Lo´pez, M.-T.; Goza´lez Mun˜iz, R. Tetrahedron
1993, 49, 8911.
(40) J ohnson, C. R.; Golebiowski, A.; Sundram, H.; Miller, M. W.;
Dwaihy, R. L. Tetrahedron Lett. 1994, 36, 653.
(41) Humeres, E.; Debacher, N. A.; Sierra, M. M. de S.; Franco, J .
D.; Schutz, A. J . Org. Chem. 1998, 63, 1598.

7610 J . Org. Chem., Vol. 66, No. 23, 2001
Garc´ıa-Moreno et al.
Exp er im en ta l Section
binding subsite. Introduction of aglycon-mimicking groups
may then result in stronger inhibition.
Gen er a l P r oced u r es. Optical rotations were measured at
room temperature in 1 cm or 1 dm tubes. IR spectra were
recorded on a FT-IR instrument. 1H (and 13C) NMR spectra
were recorded at 500 (125.7) and 300 (75.5) MHz. In the
FABMS spectra, the primary beam consisted of Xe atoms with
a maximum energy of 8 keV. The samples were dissolved in
m-nitrobenzyl alcohol or thioglycerol as the matrixes, and the
positive ions were separated and accelerated over a potential
of 7 keV. NaI was added as a cationizing agent. In the CIMS
spectra, isobutane was used as the reactive gas (500 mA, 8
kV). Visualization of TLC plates was effected by UV light and
by charring with 10% sulfuric acid or 0.2% w/v cerium(IV)
sulfate-5% ammonium molybdate in 2 M H2SO4. Fully
unprotected compounds were purified by GPC (Sephadex G-10,
1:1 MeOH-H2O). Acetylations were effected conventionally
with 1:1 pyridine-Ac2O (10 mL/1 g of sample). Deacetylations
were effected by treatment with methanolic NaOMe (0.1 equiv/
mol of acetate) at room temperature for 3 h, followed by
neutralization with Amberlite IR-120 (H+) ion-exchange resin.
Microanalyses were performed by the Instituto de Investiga-
ciones Qu´ımicas (Sevilla, Spain).
Ma t er ia ls. 5-Amino-5-deoxy-1,2-O-isopropylidene-â-L-id-
ofuranose (7) was prepared from commercial D-glucofuranu-
rono 6,3-lactone according to the literature.18a,22 2,3,4,6-Tetra-
O-acetyl-â-D-glucopyranosyl isothiocyanate (8) was synthesized
from the corresponding per-O-acetyl glucopyranosyl bromide
by treatment with potassium thiocyanate and tetra-n-buty-
lammonium hydrogensulfate in acetonitrile, following the
procedure of Camarasa et al.23 Methyl 2,3,4-tri-O-acetyl-6-
deoxy-6-isothiocyanato-R-D-glucopyranoside (9) was obtained
by isothiocyanation of the corresponding 6-amino-6-deoxysugar
using thiophosgene as reported.24 3,6-Di-O-acetyl-5-azido-5-
deoxy-1,2-O-isopropylidene-â-L-idofuranose (16) was obtained
by conventional acetylation of 5-azido-5-deoxy-1,2-O-isopro-
pylidene-â-L-idofuranose18a,22 in quantitative yield (see Sup-
porting Information). 5-Azido-5-deoxy-1,2-O-isopropylidene-R-
D-xylofuranose (36) was prepared from the corresponding 5-O-
tosyl derivative by treatment with sodium azide.35 TFA-
catalyzed hydrolysis of the acetonide protecting group of 36
and further conventional acetylation afforded the triacetate
37 as a mixture of the R- and â-anomers (Scheme 5; see
Supporting Information). 9-Fluorenylmethyloxycarbonyl isothio-
cyanate was prepared from commercial chloride by reaction
with potassium isothiocyanate.44 The R-glucosidase (from
brewer yeast), â-glucosidase (from almonds), â-glucosidase
(from bovine liver, cytosolic), and R-galactosidase (from green
coffee beans) used in the inhibition studies as well as the
corresponding p-nitrophenyl glycoside substrates were pur-
chased from Sigma Chemical Company.
Reagents and solvents were commercial grade and were
used as supplied, with the following exceptions: potassium
thiocyanate was dried with heating under vacuum at 80 °C,
DMF was distilled from BaO, methanol was distilled from
methylmagnesium iodide, pyridine was distilled from KOH,
and acetic anhydride was distilled from freshly melted sodium
acetate.
Gen er a l P r oced u r e for th e In h ibition Assa y. Inhibitory
potencies were determined by spectrophotometrically measur-
ing the residual hydrolytic activities of the glycosidases against
the respective p-nitrophenyl R- or â-D-glucopyranoside or
p-nitrophenyl R-D-galactopyranoside in the presence of the
corresponding 6-oxacalystegine derivative. Each assay was
performed in phosphate buffer at the optimal pH for each
enzyme. The reactions were initiated by addition of the enzyme
to a solution of the substrate in the presence or absence of
various concentrations of the inhibitor. After the mixture was
incubated for 10-30 min at 37 °C, the reaction was quenched
by addition of 1 M Na2CO3. The absorbance of the resulting
mixture was determined at 400 nm. The Ki value and enzyme
inhibition mode were determined from the slope of Lin-
The high selectivity and potency observed for the
oxacalystegine derivatives with aromatic substituents 25,
26, 30, and 33 in the inhibition of the mammaliam
cytosolic â-glucosidase, an enzyme known to possess a
hydrophobic binding site and for which aromatic â-D-
glucosides are good substrates,43 confirm the above
assumptions and strongly support a 1-azasugar mode of
action for calystegine-type inhibitors against â-glucosi-
dases. The reverse selectivity of the N-(thio)carbamoyl-
6-oxacalystegines toward cytosolic and almond â-glucosi-
dases compared with that of calystegine B2 or the
unsubstituted 6-oxacalystegine 35, incorporating a basic
nitrogen atom, probably reflects the need for a higher
contribution of electrostatic interactions to achieve an
efficient binding in the case of the later enzyme.
The 10-fold increase in the cytosolic â-glucosidase
inhibition potency for thiocarbamoyl (e.g., 26) compared
with that for carbamoyl nor-tropane derivatives (e.g., 30)
is probably due to the higher positive charge density of
the thiourea N-atom compared with that of urea. In any
case, the rigid bicyclic structure seems to be a major
requirement for â-glucosidase inhibition. Thus, replace-
ment of the oxazolidine ring (e.g., in 26 or 30) by an open
aminoacetal center (e.g., in the xylonojirimycin analogues
49 and 52) totally abolished inhibition in both series.
No significant influence of the nature of the pseudoa-
glyconic substituent on the interaction of 6-oxacalyste-
gines with R-galactosidase was observed. The strong
decrease in inhibition potency for the unsubstituted
derivative 35 compared to that for calystegine B2, more
than 100-fold, indicates that the bridgehead hydroxy
group of 1 is actively involved in strong binding. In
contrast to the above comments for the mammalian
â-glucosidase, disrupting the cyclic aminoacetal group
has only a weak effect (a ca. 3-fold decrease) on the
inhibition potency.
Further work is necessary to determine the exact mode
of interaction of calystegine-type azasugars with glycosi-
dases. Kinetic studies with the new oxacalystegine
derivatives against both cytosolic â-glucosidase and R-ga-
lactosidase indicated fast and competitive inhibition, as
reported for the natural alkaloids. The C-1-C-5 two-atom
bridge seems to prevent recognition by R-glucosidase and
forces a recognition mode analogous to that of 1-azasug-
ars for â-glucosidase, the N-substituent interacting with
the aglycon binding subsite. In any case, our work
provides a new tool for preparing selective â-glucosidase
inhibitors of the calystegine family by exploiting the
reactivity of N-(thio)carbamoyl aminoketalic systems. A
variety of structures become available by acting on the
configuration of the monosaccharide template and the
nature of the N-substituent.
(42) Theoretical and experimental studies indicate that true transi-
tion state analogues must be essentially neutral or zwitterionic at
physiological pH to keep specificity against the target enzyme. See:
(a) J eong, J .-H.; Murray, B. W.; Takayama, S.; Wong, C.-H. J . Am.
Chem. Soc. 1996, 118, 4227. (b) Legler, G.; Finken, M.-T. Carbohydr.
Res. 1996, 292, 103. (c) Ble´riot, Y.; Genre-Grandpierre, A.; Imberty,
A.; Tellier, C. J . Carbohydr. Chem. 1996, 15, 985. (d) Deng, H.; Chan,
A. W.-Y.; Bagdasianan, C. K.; Estupin˜a´n, B.; Ganem, B.; Callender,
R. H.; Scharam, V. L. Biochemistry 1996, 35, 6037. (e) Ernert, P.;
Vasella, A.; Weber, M.; Rupitz, K.; Withers, S. G. Carbohydr. Res. 1993,
250, 113.
(43) Legler, G.; Bieberich, E. Arch. Biochem. Biophys. 1988, 260,
427.
(44) Kearney, P. C.; Fernandez, M.; Flygare, J . A. J . Org. Chem.
1998, 63, 196.

Calystegine B2 Analogues
J . Org. Chem., Vol. 66, No. 23, 2001 7611
eweaver-Burk plots and double-reciprocal analyses using a
5-De oxy-1,2-O-isop r op ylid e n e -5-[3-(2,3,4,6-t e t r a -O-
a cet yl-â-D-glu cop yr a n osyl)t h iou r eid o]-â-L-id ofu r a n ose
(15): yield 1.29 g (93%); Rf (20:1 CH2Cl2-MeOH) 0.22; [R]D
-2.6 (c 0.76, CH2Cl2); UV (MeOH) 248 nm (ꢀmM 14.5); IR (KBr)
Sigma Plot program (version 4.14, J andel Scientific).
Gen er a l P r oced u r e for th e P r ep a r a tion of 5-Deoxy-5-
th iou r eido-L-idofu r an oses (10-16). To a solution of 5-amino-
5-deoxy-1,2-O-isopropylidene-â-L-idofuranose 7 (0.5 g, 2.29
mmol) in pyridine (15 mL) were added the corresponding
9-fluorenyloxycarbonyl, methyl, benzyl, phenyl, or sugar-
derived isothiocyanate 8 or 9 (2.29 mmol) and Et3N (0.05 mL).
The mixture was stirred at room temperature for 3-24 h (TLC)
and concentrated. The resulting residue was chromatographed
with the indicated eluent to give the thiourea adduct 10-13,
15, or 16, respectively, as an amorphous solid. The N-
monosubstituted thiourea 14 was obtained from the N′-Fmoc-
protected derivative 10 (195 mg, 0.39 mmol) by treatment with
20% methanolic piperidine (1 mL) in CH2Cl2 (1.6 mL) at room
temperature for 2 h and further purification by column
chromatography (45:5:3 AcOEt-EtOH-H2O).
νmax 3428, 2951, 1760, 1553 cm-1 1H NMR (300 MHz,
;
CD3OD, 313 K) (see also Table 2, Supporting Information) δ
2.02, 2.00 (6 H), 1.97 (4 Ac), 1.44, 1.28 (2 s, each 3 H, Me2C);
13C NMR (75.5 MHz, CD3OD) δ 185.3, 172.3, 171.6, 171.5,
171.3, 112.6, 105.9, 86.9, 83.5, 80.4, 75.6, 74.8, 74.4, 71.9, 69.7,
63.1, 61.8, 56.5, 27.0, 26.3, 20.8-20.4; FABMS m/z 631 (100,
[M + Na]+). Anal. Calcd for C24H36N2O14S: C, 47.36; H, 5.97;
N, 4.60; S, 5.27. Found: C, 47.01; H, 5.90; N, 4.33; S, 5.00.
5-Deoxy-1,2-O-isop r op ylid en e-5-[3-(m eth yl 2,3,4-tr i-O-
a cetyl-6-d eoxy-r-D-glu cop yr a n osyd -6-yl)th iou r eid o]-â-L-
id ofu r a n ose (16): yield 1.0 g (76%); Rf (20:1 CH2Cl2-MeOH)
0.26; [R]D +62.0 (c 1.0, CH2Cl2); UV (CH2Cl2) 248 nm (ꢀmM 11.7);
IR (KBr) νmax 3497, 2961, 1750, 1562 cm-1; 1H NMR (500 MHz,
CDCl3, 323 K) (see also Table 2, Supporting Information) δ
6.82 (bd, 1 H, NH′), 6.50 (bs, 1 H, NH), 3.38 (s, 3 H, OMe),
2.05, 2.00, 1.96 (3 s, each 3 H, 3 Ac), 1.46, 1.29 (2 s, each 3 H,
Me2C); 13C NMR (125.7 MHz, CDCl3) δ 183.4, 170.6-169.9,
111.7, 104.4, 96.5, 84.7, 80.0, 75.1, 70.7, 69.5, 68.8, 68.0, 63.3,
55.4, 54.9, 44.4, 26.6, 25.9, 20.9-20.5; FABMS m/z 603 (100,
[M + Na]+). Anal. Calcd for C23H36N2O13S: C, 47.58; H, 6.25;
N, 4.83. Found: C, 47.41; H, 6.23; N, 4.66.
Gen er a l P r oced u r e for th e P r ep a r a tion of 5-Ca r bod i-
im id o-5-d eoxy-L-id ofu r a n oses (18-20). A solution of azide
17 (1.0 g, 3.04 mmol) in toluene (45 mL) was stirred under
nitrogen for 30 min. Then, the corresponding phenyl or sugar-
derived isothiocyanate 8 or 9 (3.04 mmol) and a solution of
TPP (0.88 g, 3.34 mmol) in toluene (20 mL) were added
dropwise at room temperature. The reaction mixture was
stirred for 24 h and concentrated, and the residue was purified
by column chromatography (1:3 f 1:1 EtOAc-petroleum
ether). The resulting carbodiimides were isolated as amor-
phous solids.
5-Deoxy-5-[3-(9-flu or en ylm eth yloxyca r bon yl)th iou r e-
id o]-1,2-O-isop r op ylid en e-â-L-id ofu r a n ose (10): yield 0.92
g (80%); Rf (2:1 EtOAc-petroleum ether) 0.53; [R]D -18.4 (c
1.0, CH2Cl2); UV (CH2Cl2) 266 nm (ꢀmM 36.0); IR (KBr) νmax
3275, 2986, 1723, 1537 cm-1; 1H NMR (500 MHz, CDCl3) (see
also Table 2, Supporting Information) δ 10.11 (d, 1 H, J NH,5
)
8.0 Hz, NH), 8.58 (s, 1 H, NH), 7.74-7.30 (m, 8 H, aromatic),
2
3
4.45 (dd, 1 H, J H,H ) 10.6 Hz, J H,H ) 6.8 Hz, CH2), 4.42 (dd,
1 H, CH2), 4.20 (t, 1 H, CH), 1.48, 1.28 (2 s, each 3 H, Me2C);
13C NMR (125.7 MHz, CDCl3) δ 179.1, 152.4, 142.9, 141.2,
127.9, 127.2, 124.9, 120.1, 111.8, 104.7, 85.1, 79.2, 75.3, 68.3,
62.0, 55.7, 46.5, 26.8, 26.1; CIMS m/z 501 (50, [M + H]+). Anal.
Calcd for C25H28N2O7S: C, 59.99; H, 5.64; N, 5.59. Found: C,
59.80; H, 5.56; N, 5.53.
5-Deoxy-1,2-O-isop r op ylid en e-5-(3-m eth ylth iou r eid o)-
â-L-id ofu r a n ose (11): yield 0.58 g (87%); Rf (20:1 CH2Cl2-
MeOH) 0.27; [R]D -19.5 (c 1.1, MeOH); UV (MeOH) 240 nm
1
(ꢀmM 9.1); IR (KBr) νmax 3376, 2926, 1572 cm-1; H NMR (300
MHz, CD3OD, 313 K) (see also Table 2, Supporting Informa-
tion) δ 2.97 (s, 3 H, MeNH), 1.45, 1.30 (2 s, each 3 H, Me2C);
13C NMR (75.5 MHz, CD3OD, 313 K) δ 182.5, 112.6, 105.8,
86.8, 81.0, 75.6, 62.5, 56.5, 33.0, 26.9, 26.3; FABMS m/z 315
(100, [M + Na]+). Anal. Calcd for C11H20N2O5S: C, 45.19; H,
6.90; N, 9.58; S, 10.79. Found: C, 45.20; H, 6.72; N, 9.53; S,
11.07.
3,6-Di-O-acetyl-5-deoxy-1,2-O-isopr opyliden e-5-(3-ph en -
ylca r bod iim id o)-â-L-id ofu r a n ose (18): yield 0.98 g (80%);
Rf (2:1 EtOAc-petroleum ether) 0.83; [R]D -91.4 (c 0.5,
CH2Cl2); IR (KBr) νmax 2991, 2139, 1750, 1593, 1501 cm-1; 1H
NMR (300 MHz, CDCl3) (see also Table 2, Supporting Infor-
mation) δ 7.25-7.03 (m, 5 H, Ph), 2.08, 1.94 (2 s, each 3 H, 2
Ac), 1.46, 1.25 (2 s, each 3 H, Me2C); 13C NMR (75.5 MHz,
CDCl3) δ 170.2, 169.7, 139.3, 137.8, 129.3, 125.2, 125.1, 123.9,
112.3, 104.3, 83.5, 78.2, 75.9, 64.3, 55.6, 26.6, 26.1, 20.7, 20.5;
FABMS m/z 427 (35, [M + Na]+). Anal. Calcd for C20H24N2O7:
C, 59.40; H, 5.94; N, 6.93. Found: C, 59.35; H, 6.11; N, 6.91.
3,6-Di-O-a ce t yl-5-d e oxy-1,2-O-isop r op ylid e n e -5-[3-
(2,3,4,6-tetr a-O-acetyl-â-D-glu copyr an osyl)car bodiim ido]-
â-L-id ofu r a n ose (19): yield 1.81 g (90%); Rf (1:1 EtOAc-
petroleum ether) 0.34; [R]D -15.4 (c 1.0, CH2Cl2); IR (KBr) νmax
2963, 2143, 1750, 1554 cm-1; 1H NMR (300 MHz, CDCl3) (see
also Table 2, Supporting Information) δ 2.15-1.98 (6 s, each
3 H, 6 Ac), 1.52, 1.31 (2 s, each 3 H, Me2C); 13C NMR (75.5
MHz, CDCl3) δ 170.5-169.2, 138.3, 112.3, 104.1, 84.5, 83.6,
78.2, 75.7, 73.6, 72.9, 72.4, 68.1, 63.9, 61.9, 54.9, 26.6, 26.1,
20.7-20.5; FABMS m/z 681 (100, [M + Na]+). Anal. Calcd for
5-(3-Ben zylth iou r eid o)-5-d eoxy-1,2-O-isop r op ylid en e-
â-L-id ofu r a n ose (12): yield 0.51 g (60%); Rf (20:1 CH2Cl2-
MeOH) 0.26; [R]D -27.7 (c 1.0, MeOH); UV (MeOH) 242 nm
(ꢀmM 19.6); IR (KBr) νmax 3395, 2980, 1562, 1537 cm-1; 1H NMR
(300 MHz, CD3OD, 318 K) (see also Table 2, Supporting
Information) δ 7.32-7.19 (m, 5 H, Ph), 4.49 (s, 2 H, CH2), 1.44,
1.28 (2 s, each 3 H, Me2C); 13C NMR (75.5 MHz, CD3OD, 318
K) δ 184.2, 139.8, 129.5, 128.6, 128.2, 112.8, 105.9, 86.9, 81.1,
75.8, 62.6, 56.3, 49.0, 27.0, 26.4; FABMS m/z 391 (100, [M +
Na]+). Anal. Calcd for C17H24N2O5S: C, 55.42; H, 6.56; N, 7.60.
Found: C, 55.08; H, 6.74; N, 7.54.
5-Deoxy-1,2-O-isop r op ylid en e-5-(3-p h en ylth iou r eid o)-
â-L-id ofu r a n ose (13): yield 0.73 g (90%); Rf (20:1 CH2Cl2-
MeOH) 0.24; [R]D +14.6 (c 0.69, MeOH); UV (MeOH) 250 nm
(ꢀmM 14.4); IR (KBr) νmax 3428, 2959, 1657 cm-1; 1H NMR (300
MHz, CD3OD, 313 K) (see also Table 2, Supporting Informa-
tion) δ 7.45-7.15 (m, 5 H, Ph), 1.44, 1.29 (2 s, each 3 H, Me2C);
13C NMR (75.5 MHz, CD3OD, 313 K) δ 182.0, 139.6, 130.2,
126.7, 125.3, 112.8, 105.9, 86.9, 80.6, 75.8, 62.4, 56.6, 27.0, 26.4;
C
28H38N2O16: C, 51.06; H, 5.82; N, 4.25. Found: C, 50.91; H,
5.74; N, 4.10.
3,6-Di-O-acetyl-5-deoxy-1,2-O-isopr opyliden e-5-[3-(m eth -
yl 2,3,4-t r i-O-a cet yl-6-d eoxy-â-D-glu cop yr a n osyd -6-yl)-
ca r bod iim id o]-â-L-id ofu r a n ose (20): yield 0.59 g (31%); Rf
(1:1 EtOAc-petroleum ether) 0.38; [R]D +51.5 (c 1.0, CH2Cl2);
IR (KBr) νmax 2961, 2137, 1742, 1574, 1514 cm-1; 1H NMR (500
MHz, CDCl3) (see also Table 3, Supporting Information) δ 3.36
(s, 3 H, OMe), 2.04-192 (5 s, each 3 H, 5 Ac), 1.44, 1.23 (2 s,
each 3 H, Me2C); 13C NMR (75.5 MHz, CDCl3) δ 170.2-169.4
(5 CO), 140.2, 112.1, 103.4, 96.5, 83.4, 78.4, 75.6, 70.5, 69.8,
69.5, 67.9, 64.1, 55.3, 54.6, 46.5, 26.4, 26.0, 20.5-20.4; FABMS
m/z 653 (100, [M + Na]+). Anal. Calcd for C27H38N2O15: C,
51.42; H, 6.07; N, 4.44. Found: C, 51.21; H, 5.77; N, 4.33.
Gen er a l P r oced u r e for th e P r ep a r a tion of 5-Deoxy-5-
u r eid o-L-id ofu r a n oses (21-23). To a solution of the corre-
sponding carbodiimide 18-20 (1.52 mmol) in 2:1 acetone-
water (45 mL) was added TFA (0.5 mL). The reaction mixture
FABMS m/z 377 (100, [M
16H22N2O5S: C, 54.22; H, 6.26; N, 7.90; S, 9.05. Found: C,
54.24; H, 6.40; N, 7.89; S, 9.05.
+
Na]+). Anal. Calcd for
C
5-Deoxy-1,2-O-isopr opyliden e-5-th iou r eido-â-L-idofu r a-
n ose (14): yield 65 mg (60%); Rf (45:5:3 AcOEt-EtOH-H2O)
0.54; [R]D -27.9 (c 0.88, MeOH); UV (MeOH) 242 nm (ꢀmM
16.2); IR (KBr) νmax 3434, 3324, 2980, 1603 cm-1; 1H NMR (300
MHz, CD3OD, 313 K) (see also Table 2, Supporting Informa-
tion) δ 1.50, 1.29 (2 s, each 3 H, Me2C); 13C NMR (75.5 MHz,
CD3OD, 313 K) δ 180.0, 113.6, 106.7, 87.8, 81.9, 76.5, 63.3,
57.8, 27.9, 27.3; CIMS m/z 279 (100, [M + H]+). Anal. Calcd
for C16H22N2O5S: C, 54.22; H, 6.26; N, 7.90; S, 9.05. Found:
C, 54.24; H, 6.40; N, 7.89; S, 9.05.

7612 J . Org. Chem., Vol. 66, No. 23, 2001
Garc´ıa-Moreno et al.
was stirred at room temperature for 2-12 h until the starting
material disappeared (TLC), and the solution was then
concentrated. Column chromatography of the residue (1:1 f
2:1 EtOAc-petroleum ether) afforded the urea as an amor-
phous solid.
2 H, CH2); 13C NMR (75.5 MHz, D2O) δ 178.4, 138.0, 128.9,
127.5, 127.1, 88.5, 74.9, 73.8, 70.6, 66.3, 58.4, 47.4; FABMS
m/z 311 (80, [M + H]+). Anal. Calcd for C14H18N2O4S: C, 50.89;
H, 5.49; N, 8.48. Found: C, 50.78; H, 5.40; N, 8.29.
(1S,2R,3S,4R,5R)-2,3,4-Tr ih ydr oxy-6-oxa-N-(N′-ph en ylth -
ioca r ba m oyl)-n or -t r op a n e (26): yield 102 mg (69%); [R]D
+66.1 (c 0.6, MeOH); Rf (45:5:3 EtOAc-EtOH-H2O) 0.55; UV
(MeOH) 245 nm (ꢀmM 13.2); IR (KBr) νmax 3341, 2928, 1537
3,6-Di-O-acetyl-5-deoxy-1,2-O-isopr opyliden e-5-(3-ph en -
ylu r eid o)-â-L-id ofu r a n ose (21): yield 0.64 g (100%); Rf (2:1
EtOAc-petroleum ether) 0.60; [R]D -2.3 (c 1.0, CH2Cl2); IR
(KBr) νmax 3384, 2996, 1750 cm-1; 1H NMR (300 MHz, CDCl3)
(see also Table 2, Supporting Information) δ 7.32-7.26 (m, Ph),
6.58 (bs, 1 H, NH′), 5.19 (d, 1 H, J NH,5 ) 8.0 Hz, NH), 2.06 (2
s, each 3 H, 2 Ac), 1.51, 1.31 (2 s, each 3 H, Me2C); 13C NMR
(75.5 MHz, CDCl3) δ 170.8, 169.8, 154.7, 138.3, 129.1, 123.7,
120.3, 112.7, 104.3, 83.5, 77.1, 64.5, 47.6, 26.6, 20.7; FABMS
m/z 445 (100, [M + Na]+). Anal. Calcd for C20H26N2O8: C,
56.87; H, 6.16; N, 6.63. Found: C, 56.68; H, 6.43; N, 6.60.
3,6-Di-O-acetyl-5-deoxy-1,2-O-isopr opyliden e-5-[(2,3,4,6-
t et r a -O-a cet yl-â-D-glu cop yr a n osyl)u r eid o]-â-L-id ofu r a -
n ose (22): yield 0.67 g (65%); Rf (3:1 EtOAc-petroleum ether)
0.34; [R]D -2.8 (c 1.0, CH2Cl2); IR (KBr) νmax 3393, 2986, 1750,
1696, 1547 cm-1; 1H NMR (500 MHz, CDCl3, 313 K) (see also
Table 2, Supporting Information) δ 5.43 (d, 1 H, J N′H,1′ ) 9.3
Hz, NH), 5.11 (d, 1 H, J NH,5 ) 7.1 Hz, NH), 2.05-1.95 (6 s,
each 3 H, 6 Ac), 1.48, 1.27 (2 s, each 3 H, Me2C); 13C NMR
(75.5 MHz, CDCl3) δ 170.8-169.5, 155.7, 112.1, 104.2, 83.4,
80.1, 77.1, 76.2, 72.9 (2 C), 68.1 (2 C), 63.9, 61.6, 26.4, 26.0,
20.6-20.4; FABMS m/z 699 (100, [M + Na]+). Anal. Calcd for
cm-1 1H NMR (500 MHz, D2O, 323 K) (see also Table 3,
;
Supporting Information) δ 7.85-7.65 (m, 5 H, PhN); 13C NMR
(125.5 MHz, D2O, 313 K) δ 179.3, 139.7, 130.4, 128.6, 128.3,
89.7, 76.1, 74.8, 71.7, 67.4, 59.9; FABMS m/z 319 (40, [M +
Na]+). Anal. Calcd for C13H16N2O4S: C, 52.69; H, 5.44; N, 9.45.
Found: C, 52.58; H, 5.33; N, 9.46.
(1S,2R,3S,4R,5R)-2,3,4-Tr ih ydr oxy-6-oxa-N-th iocar bam -
oyl-n or -tr op a n e (27): column chromatography, eluent 6:1
CH2Cl2-MeOH; yield 106 mg (96%); Rf (4:1 CH2Cl2-MeOH)
0.35; [R]D +58.0 (c 1.0, H2O); UV (MeOH) 250 nm (ꢀmM 13.0);
IR (KBr) νmax 3407, 3380, 1656, 1553 cm-1; 1H NMR (300 MHz,
D2O, 313 K) (see also Table 3, Supporting Information); 13C
NMR (75.5 MHz, D2O, 313 K) δ 177.5, 88.5, 75.2, 73.9, 70.7,
66.9, 58.8; FABMS m/z 221 (30, [M + H]+). Anal. Calcd for
C7H12N2O4S: C, 38.17; H, 5.49; N, 12.72. Found: C, 38.39; H,
5.39; N, 12.69.
(1S,2R,3S,4R,5R)-N-[N′-(â-D-Glu cop yr a n osyl)t h ioca r -
b a m oyl]-2,3,4-t r ih yd r oxy-6-oxa -n or -t r op a n e (28): yield
124 mg (65%); [R]D +21.7 (c 0.65, MeOH); Rf (6:3:1 MeCN-
H2O-NH4OH) 0.33; UV (MeOH) 253 nm (ꢀmM 7.0); IR (KBr)
C
28H40N2O17: C, 49.70; H, 5.96; N, 4.14. Found: C, 49.41; H,
5.87; N, 4.14.
ν
max 3404, 1657, 1553 cm-1; 1H NMR (500 MHz, D2O) (see also
3,6-Di-O-acetyl-5-deoxy-1,2-O-isopr opyliden e-5-[3-(m eth -
yl 2,3,4-t r i-O-a cet yl-6-d eoxy-â-D-glu cop yr a n osyd -6-yl)-
u r eid o]-â-L-id ofu r a n ose (23): yield 0.95 g (96%); Rf (3:1
EtOAc-petroleum ether) 0.25; [R]D +57.8 (c 1.0, CH2Cl2); IR
Table 3, Supporting Information); 13C NMR (125.7 MHz, D2O)
δ 179.2, 88.0, 84.1, 77.0, 76.1, 74.3, 72.9, 71.2, 69.8, 68.7, 65.9,
60.0, 57.7; FABMS m/z 405 (40, [M + Na]+). Anal. Calcd for
C
13H22N2O9S: C, 40.83; H, 5.80; N, 7.32. Found: C, 40.81; H,
1
(KBr) νmax 3387, 2988, 1750, 1651, 1559 cm-1; H NMR (500
5.52; N, 7.15.
MHz, CDCl3) (see also Table 2, Supporting Information) δ 4.90
(m, 2 H, 2 NH), 3.38 (s, 3 H, OMe), 2.07-1.98 (5 s, each 3 H,
5 Ac), 1.49, 1.29 (2 s, each 3 H, Me2C); 13C NMR (125.5 MHz,
CDCl3) δ 170.6-169.8, 157.1, 112.2, 104.2, 96.8, 83.6, 77.6,
76.4, 71.0, 70.0, 69.4, 68.2, 64.4, 55.3, 47.9, 40.3, 26.5, 26.1,
20.6-20.5; FABMS m/z 671 (100 [M + Na]+). Anal. Calcd for
(1S,2R,3S,4R,5R)-2,3,4-Trihydroxy-N-[N′-(methyl6-deoxy-
r-D-glu cop yr a n osyd -6-yl)th ioca r ba m oyl]-6-oxa -n or -tr o-
p a n e (29): column chromatography, eluent 20:1 f 5:1 EtOAc-
EtOH; yield 176 mg (89%); [R]D +85.0 (c 1.1, MeOH); Rf (1:1
EtOAc-EtOH) 0.36; UV (MeOH) 249 nm (ꢀmM 13.8); IR (KBr)
ν
max 3362, 1651, 1539 cm-1; 1H NMR (500 MHz, D2O) (see also
C
27H40N2O16: C, 49.99; H, 6.22; N, 4.32. Found: C, 50.07; H,
Table 3, Supporting Information) δ 3.27 (s, 3 H, OMe); 13C
NMR (125.7 MHz, D2O) δ 178.4, 99.3, 88.6, 75.1, 73.9, 73.1,
71.8, 71.5, 70.7, 69.7, 66.3, 58.4, 55.9, 46.2; FABMS m/z 603
(40, [M + Na]+). Anal. Calcd for C14H24N2O9S: C, 42.42; H,
6.10; N, 7.07. Found: C, 42.31; H, 5.99; N, 6.93.
6.30; N, 4.34.
Gen er a l P r oced u r e for th e P r ep a r a tion of N-(Th io)-
ca r ba m oyl-6-oxa ca lystegin es (24-32). A solution of 11-
14 or of the product of deacetylation (NaOMe/MeOH) of 15,
16, or 21-23 (0.5 mmol) in 90% TFA-H2O (5 mL) was stirred
at room temperature for 20-75 min until the starting material
disappeared (TLC, 45:5:3 EtOAc-EtOH-H2O). The reaction
mixture was concentrated and the residue coevaporated
several times with water. An aqueous solution was further
neutralized with Amberlite IRA-68 (OH-) ion-exchange resin,
filtered, and freeze-dried. Where stated, the residue was
purified by column chromatography using the eluent indicated
in each case. In all cases, the fully unprotected compounds
were subjected to GPC (Sephadex G-10, 1:1 MeOH-H2O) to
afford the target 6-oxacalystegine derivatives as white foams
after lyophilization of an aqueous solution.
(1S,2R,3S,4R,5R)-2,3,4-Tr ih yd r oxy-6-oxa -N-(N′-p h en yl-
ca r ba m oyl)-n or -tr op a n e (30): column chromatography, elu-
ent 15:1 MeCN-H2O; yield 112 mg (80%); [R]D +97.4 (c 1.0,
H2O); Rf (15:1 MeCN-H2O) 0.60; IR (KBr) νmax 3333, 1672,
1529 cm-1 1H NMR (500 MHz, D2O) (see also Table 3,
;
Supporting Information) δ 7.34-7.10 (m, 5 H, Ph); 13C NMR
(75.5 MHz, D2O) δ 157.9, 139.5, 131.2, 127.0, 124.2, 87.3, 75.4,
74.9, 71.9, 65.4, 57.3; FABMS m/z 303 (100, [M + Na]+). Anal.
Calcd for C13H14O5N2: C, 55.71; H, 5.75; N, 9.99. Found: C,
55.48; H, 5.60; N, 9.79.
(1S,2R,3S,4R,5R)-N-[N′-(â-D-Glu copyr an osyl)car bam oyl]-
2,3,4-tr ih yd r oxy-6-oxa -n or -tr op a n e (31): column chroma-
tography, eluent 15:1 MeCN-H2O; yield 174 mg (95%); [R]D
+25.0 (c 1.0, H2O); Rf (6:3:1 MeCN-H2O-NH4OH) 0.30; IR
(1S,2R,3S,4R,5R)-2,3,4-Tr ih yd r oxy-N-(N′-m et h ylt h io-
ca r ba m oyl)-6-oxa -n or -tr op a n e (24): column chromatogra-
phy, eluent 45:5:3 AcOEt-EtOH-H2O; yield 76 mg (65%); [R]D
+47.7 (c 0.6, H2O); Rf (45:5:3 EtOAc-EtOH-H2O) 0.39; UV
1
(KBr) νmax 3385, 2907, 1651, 1541 cm-1; H NMR (500 MHz,
D2O) (see also Table 3, Supporting Information); 13C NMR
(125.7 MHz, D2O) δ 156.1, 85.7, 80.8, 76.8, 76.1, 74.7, 74.0,
71.1, 70.8, 68.9, 64.9, 60.2, 56.3; FABMS m/z 389 (100, [M +
Na]+). Anal. Calcd for C13H22N2O10: C, 42.68; H, 6.05; N, 7.65.
Found: C, 42.42; H, 5.97; N, 7.41.
(MeOH) 246 nm (ꢀmM 9.4); IR (KBr) νmax 3364, 2912, 1553 cm-1
;
1H NMR (500 MHz, D2O, 323 K) (see also Table 3, Supporting
Information) δ 3.52 (s, 3 H, MeNH); 13C NMR (75.5 MHz, D2O,
323 K) δ 177.9, 88.4, 75.0, 73.8, 70.5, 67.4, 58.2, 31.9; FABMS
m/z 257 (80, [M + Na]+). Anal. Calcd for C8H14N2O4S: C, 41.02;
H, 5.98; N, 11.96; S, 13.68. Found: C, 40.82; H, 5.94; N, 11.93;
S, 13.79.
(1S,2R,3S,4R,5R)-2,3,4-Trihydroxy-N-[N′-(methyl6-deoxy-
r-D -g lu c o p y r a n o s y d -6-y l)c a r b a m o y l]-6-o x a -n or -t r o -
p a n e (32): column chromatography, eluent 7:1 f 5:1 MeCN-
H2O; yield 162 mg (85%); [R]D +61.2 (c 1.0, H2O); Rf (3:1
(1S,2R,3S,4R,5R)-N-(N′-Ben zylth ioca r ba m oyl)-2,3,4-tr i-
h yd r oxy-6-oxa -n or -tr op a n e (25): column chromatography,
eluent 3:1 EtOAc-petroleum ether; yield 93 mg (60%): Rf (45:
5:3 EtOAc-EtOH-H2O) 0.46; [R]D +63.1 (c 1.0, MeOH); UV
(MeOH) 250 nm (ꢀmM 14.7); IR (KBr) νmax 3344, 2948, 1605,
MeCN-H2O) 0.55; IR (KBr) νmax 3380, 2910, 1650, 1543 cm-1
;
1H NMR (500 MHz, D2O) (see also Table 3, Supporting
Information) δ 3.26 (s, 3 H, OMe); 13C NMR (125.7 MHz, D2O)
δ 157.9, 99.1, 87.1, 75.2, 74.7, 73.0, 71.6, 71.4, 71.3, 70.3, 65.1,
57.0, 54.8, 41.4; FABMS m/z 381 (40, [M + Na]+). Anal. Calcd
1535 cm-1 1H NMR (300 MHz, D2O) (see also Table 3,
;
Supporting Information) δ 7.40-7.30 (m, 5 H, PhN), 4.70 (bs,

Calystegine B2 Analogues
J . Org. Chem., Vol. 66, No. 23, 2001 7613
for C14H24N2O10: C, 42.21; H, 6.36; N, 7.36. Found: C, 42.20;
H, 6.16; N, 7.37.
pH 8 by addition of saturated aqueous NaHCO3. Isothiocyan-
ate 38 (140 mg, 0.44 mmol) in acetone (6 mL) was then added,
and the reaction mixture was stirred for 3 h. Acetone was
removed under reduced pressure, and the aqueous phase was
extracted with CH2Cl2 (2 × 5 mL). The combined organic phase
was dried (MgSO4) and concentrated, and the resulting residue
was chromatographed (1:2 f 1:1 EtOAc-petroleum ether) to
give 39 (114 mg, 75%) as an amorphous solid: Rf (1:1 EtOAc-
petroleum ether) 0.31; UV (CH2Cl2) 250 nm (ꢀmM 19.1); IR
5-Ben zyloxyca r b on yla m in o-5-d eoxy-1,2-O-isop r op yl-
id en e-â-L-id ofu r a n ose (33). A solution of 5-amino-5-deoxy-
1,2-O-isopropylidene-â-L-idofuranose 7 (222 mg, 1.02 mmol)
in H2O (3.46 mL) was adjusted to pH 8 by addition of solid
Na2CO3 at 0 °C, and benzyl chloroformate (0.22 mL, 1.53
mmol, 1.5 equiv) was added. The reaction mixture was stirred
at 0 °C for 10 min and then at room temperature for 2 h. The
mixture was extracted with EtOAc, and the organic phase was
dried (MgSO4), filtered, and concentrated. The resulting
residue was purified by column chromatography (20:1 CH2-
Cl2-MeOH) to give 33 (306 mg 85%) as an amorphous solid:
[R]D -25.5 (c 1.0, CH2Cl2) [lit.31 -24 (c 0.45, MeOH); lit.45 -24.5
(c 0.5, CHCl3)]; Rf (20:1 CH2Cl2-MeOH) 0.20; IR (KBr) νmax
3418, 1699, 1645, 1514 cm-1; 1H NMR (500 MHz, CDCl3) (see
also Table 3, Supporting Information) δ 7.47-7.26 (m, 5 H,
1
(KBr) νmax 3383, 2928, 1750, 1651, 1543 cm-1; H NMR (500
MHz, CDCl3, 313 K) (see also Table 4, Supporting Information)
δ 6.25 (bs, J N′H,Me ) 4.6 Hz, N′HR-anomer), 6.11 (d, J N′H,Me ) 4.5
Hz, N′Hâ-anomer), 6.03 (bt, NHR-anomer), 5.98 (bt, NHâ-anomer), 2.98
(d, MeNH), 2.17-2.07 (Ac); 13C NMR (125.7 MHz, CDCl3) δ
183.5, 170.3-169.3, 98.9, 92.9, 80.9, 80.1, 77.0, 76.2, 74.8, 44.6,
44.0, 30.5, 20.7-20.2; FABMS m/z 371 (100, [M + Na]+). Anal.
Calcd for C13H19O7N2S: C, 44.95; H, 5.51; N, 8.06. Found: C,
44.91; H, 5.43; N, 7.91.
1,2,3-Tr i-O-acetyl-5-deoxy-5-(3-ph en ylth iou r eido)-r- an d
-â-D-Xylofu r a n ose (40). A solution of aniline (51 µL, 0.5
mmol) and isothiocyanate 38 (180 mg, 0.5 mmol) in pyridine
(4 mL) was stirred for 24 h at room temperature. The solvent
was removed and the residue purified by column chromatog-
raphy (1:3 f 1:1 EtOAc-petroleum ether) to afford 40 (220
mg, 95%) as an amorphous solid: Rf (1:1 EtOAc-petroleum
ether) 0.54; UV (CH2Cl2) 264 nm (ꢀmM 15.6); IR (KBr) νmax 3335,
3109, 2963, 1748, 1667, 1530 cm-1; 1H NMR (500 MHz, CDCl3,
313 K) (see also Table 4, Supporting Information) δ 7.90 (s,
N′H), 7.30-7.18 (Ph), 6.35 (bt, 1 H, J NH,H5R ) 6.0 Hz,
NHR-anomer), 6.26 (bt, 1 H, J NH,H5â ) 9.0 Hz, NHâ-anomer), 2.08-
1.99 (Ac); 13C NMR (75.5 MHz, CDCl3) δ 181.8, 170.2-169.1,
135.4, 130.2, 127.4, 125.0, 98.7, 92.6, 79.9, 79.7, 76.1, 75.9, 74.5,
74.4, 45.0, 44.2, 20.9-20.2; FABMS m/z 433 (100, [M + Na]+).
Anal. Calcd for C18H22O7N2S: C, 52.67; H, 5.40; N, 6.83.
Found: C, 52.93; H, 5.35; N, 6.69.
2
Ph), 5.35 (sa, 1 H, NH), 5.13 (d, 2 H, J H,H ) 12.3 Hz, CH2),
2.17 (sa, 1 H, CH), 1.49, 1.31 (2 s, each 3 H, Me2C); 13C NMR
(125.7 MHz, CDCl3) δ 154.0, 135.9-128.0, 111.7, 104.5, 85.1,
80.1, 75.1, 63.3, 51.8, 26.7, 26.0; FABMS m/z 376 (100, [M +
Na]+). Anal. Calcd for C17H23O7N: C, 57.78; H, 6.56; N, 3.96.
Found: C, 57.91; H, 6.37; N, 3.76.
(1S,2R,3S,4R,5R)-N-(Ben zyloxyca r b on yl)-2,3,4-t r ih y-
d r oxy-6-oxa -n or -tr op a n e (34). A solution of 33 (300 mg, 1
mmol) was treated with 90% TFA-H2O and worked up as
described above for the preparation of 24-32. The residue was
purified by column chromatography using 20:1 f 9:1 CH2Cl2-
MeOH as the eluent to give 34 (180 mg, 80%) as a white foam
after the freeze-drying of an aqueous solution: [R]D +41.3 (c
1.0, H2O); Rf (9:1 CH2Cl2-MeOH) 0.39; IR (KBr) νmax 3389,
1
2963, 1699, 1514 cm-1; H NMR (300 MHz, D2O, 313 K) (see
also Table 3, Supporting Information) δ 7.47-7.26 (m, 5 H,
Ph), 5.47 (s, 2 H, CH2); 13C NMR (125.7 MHz, D2O, 313 K) δ
152.9, 134.2, 127.3, 127.2, 126.4, 84.7, 73.6, 73.0, 69.8, 64.4,
54.8; FABMS m/z 296 (100, [M + H]+). Anal. Calcd for
1,2,3-Tr i-O-a cetyl-5-d eoxy-5-[3-(2,3,4,6-tetr a -O-a cetyl-
â-D-glu cop yr a n osyl)th iou r eid o]-r- a n d -â-D-Xylofu r a n ose
(41). A solution of 2,3,4,6-tetra-O-acetyl-â-D-glucopyranosyl-
amine (314 mg, 0.9 mmol) and isothiocyanate 38 (285 mg, 0.9
mmol) in pyridine (6 mL) was stirred for 48 h at room
temperature. The solvent was removed and the residue puri-
fied by column chromatography (1:1 EtOAc-petroleum ether)
to yield 41 (418 mg, 70%) as an amorphous solid: Rf (2:1
EtOAc-petroleum ether) 0.54; UV (CH2Cl2) 254 nm (ꢀmM 10.3);
IR (KBr) νmax 3362, 2958, 1750, 1651, 1541 cm-1; 1H NMR (500
MHz, CDCl3, 318 K) (see also Table 4, Supporting Information)
δ 6.76 (d, J N′H,1′ ) 9.0 Hz, N′HR-anomer), 6.70 (d, J N′H,1′ ) 9.0
Hz, N′Hâ-anomer), 6.64 (m, NHR-anomer), 6.60 (dd, J NH,5a ) 6.0
Hz, J NH,5b ) 4.6 Hz, NHâ-anomer), 2.11-1.99 (Ac); 13C NMR
(125.7 MHz, CDCl3) δ 183.8, 171.0-169.1, 98.9, 92.9, 82.6, 80.3,
79.9, 77.0, 76.6, 74.6, 73.3, 72.9, 70.8, 68.2, 61.7, 43.9, 20.9-
20.2; FABMS m/z 687 (100, [M + Na]+). Anal. Calcd for
C
14H17O6N: C, 56.94; H, 5.80; N, 4.74. Found: C, 56.79; H,
5.80; N, 4.64.
(1S,2R,3S,4R,5R)-2,3,4-Tr ih yd r oxy-6-oxa -n or -t r op a n e
(35). A solution of 34 (150 mg, 0.5 mmol) in 2:1 EtOAc-MeOH
(30 mL) was hydrogenated at atmospheric pressure for 10 min
using 10% Pd/C (90 mg) as a catalyst.46 The suspension was
filtered through Celite and concentrated, and the residue was
purified by GPC (Sephadex G-10, 1:1 MeOH-H2O) to afford
35 (48 mg, 60%) as a white foam after the freeze-drying of an
aqueous solution: [R]D +103.8 (c 1.0, H2O) [lit.30a [R]D +114.5
(c 1.0, H2O)]; Rf (6:3:1 MeCN-H2O-NH4OH) 0.48; IR (KBr)
νmax 3434, 3324, 2980, 1603 cm-1 1H NMR (500 MHz, D2O,
;
313 K) (see also Table 3, Supporting Information); 13C NMR
(75.5 MHz, D2O) δ 87.9, 73.6, 73.1, 70.1, 63.3, 56.0; FABMS
m/z 162 (100, [M + H]+). Anal. Calcd for C6H11O4N: C, 44.72;
H, 6.88; N, 8.69. Found: C, 44.60; H, 6.73; N, 8.48.
C
26H36O16N2S: C, 46.98; H, 5.46; N, 4.22. Found: C, 47.03; H,
1,2,3-Tr i-O-a cetyl-5-d eoxy-5-isoth iocya n a to-r- a n d -â-
D-Xylofu r a n ose (38). To a solution of 1,2,3 tri-O-acetyl-5-
azido-5-deoxy-R- and â-D-xylofuranose 37 (400 mg, 1.33 mmol)
in dry dioxane (15 mL) were added CS2 (1.48 mL, 15 equiv)
and TPP (383 mg, 1.46 mmol) under Ar. The mixture was
stirred for 48 h at room temperature, concentrated, and
chromatographed (1:3 f 1:1 EtOAc-petroleum ether) to give
38 (400 mg, 95%) as an amorphous solid: Rf (1:1 EtOAc-
5.49; N, 4.02.
Gen er a l P r oced u r e for th e P r ep a r a tion of 5-Ca r bod i-
im id o-5-d eoxy-D-xylofu r a n oses (42-44). Aza-Wittig-type
coupling between azide 37 and the corresponding methyl,
phenyl, or glucopyranosyl isothiocyanate 8, following the
procedure described above for the synthesis of 18-20, and
purification as indicated in each case afforded the D-xylofura-
nose thioureas 42-44, respectively, as inseparable mixtures
of the R- and â-anomers.
petroleum ether) 0.60; IR (KBr) νmax 2960, 2104, 1755 cm-1
;
1H NMR (500 MHz, CDCl3) (see also Table 4, Supporting
Information) δ 2.14-2.05 (6 s, Ac); 13C NMR (125.7 MHz,
CDCl3) δ 170.1-169.1, 98.8, 92.8, 79.8, 79.4, 75.4, 75.3, 73.9,
49.5, 44.5, 44.4, 21.2-20.4; FABMS m/z 340 (100, [M + H]+).
Anal. Calcd for C12H15O7NS: C, 45.42; H, 4.76; N, 4.41.
Found: C, 45.21; H, 4.62; N, 4.34.
1,2,3-Tr i-O-a cet yl-5-d eoxy-5-(3-m et h ylca r b od iim id o)-
r- a n d -â-D-Xylofu r a n ose (42): column chromatography,
eluent 1:2 f 3:1 EtOAc-petroleum ether; yield 103 mg (45%);
Rf (1:1 EtOAc-petroleum ether) 0.53; IR (KBr) νmax 2980, 2930,
1
2139, 1748 cm-1; H NMR (500 MHz, CDCl3) (see also Table
4, Supporting Information) δ 2.96, 2.94 (2 s, MeN), 2.14, 2.13,
2.12, 2.11, 2.09, 2.08 (6 s, Ac); 13C NMR (125.7 MHz, CDCl3) δ
170.1-168.8, 140.1, 98.7, 92.7, 80.9, 80.0, 76.8, 75.5, 74.2, 73.9,
46.0, 45.5, 32.4, 21.0-20.3; CIMS m/z 315 (15, [M + H]+). Anal.
Calcd for C13H18O7N2: C, 49.72; H, 5.78; N, 8.92. Found: C,
49.47; H, 5.97; N, 8.66.
1,2,3-Tr i-O-acetyl-5-deoxy-5-(3-m eth ylth iou r eido)-r- an d
-â-D-Xylofu r a n ose (39). A solution of methylamine hydro-
chloride (119 mg, 1.76 mmol) in water (3 mL) was adjusted to
(45) Kayakiri, H.; Oku, T.; Hashimoto, M. Chem. Pharm. Bull. 1991,
39, 1397.
(46) Longer hydrogenation reaction times resulted in the formation
of 1-deoxy-L-idonojirimycin (cf. ref 31) as a side product.
1,2,3-Tr i-O-a cet yl-5-d eoxy-5-(3-p h en ylca r b od iim id o)-
r- a n d -â-D-Xylofu r a n ose (43): column chromatography,

7614 J . Org. Chem., Vol. 66, No. 23, 2001
Garc´ıa-Moreno et al.
eluent toluene f 1:5 EtOAc-toluene; yield 245 mg (87%); Rf
(1:5 EtOAc-toluene) 0.39; IR (KBr) νmax 3055, 2930, 2135,
bamoyl-2-methoxy (48-50) and 2-hydroxypiperidines 51-53,
respectively, as white foams after the freeze-drying of aqueous
solutions.
1
1751, 1589 cm-1; H NMR (500 MHz, CDCl3) (see also Table
4, Supporting Information) δ 7.28-7.07 (Ph), 2.10-2.01 (Ac);
13C NMR (125.7 MHz, CDCl3) δ 170.1-169.1, 139.3, 137.1,
129.3, 125.0, 123.8, 98.8, 92.8, 80.6, 80.1, 76.7, 75.6, 74.6, 74.3,
46.4, 45.9, 21.0-20.4; FABMS m/z 399 (100, [M + Na]+). Anal.
Calcd for C18H20O7N2: C, 57.44; H, 5.36; N, 7.44. Found: C,
57.02; H, 5.14; N, 7.61.
(2R,3R,4S,5R)-3,4,5-Tr ih yd r oxy-2-m eth oxy-N-(N′-m eth -
ylth ioca r ba m oyl)p ip er id in e (48): column chromatography,
eluents EtOAc and then 20:1 f 15:1 EtOAc-EtOH; yield 86
mg (73%); [R]D +61.1 (c 1.0, H2O); Rf (45:5:3 EtOAc-EtOH-
H2O) 0.30; UV (MeOH) 246 nm (ꢀmM 16.9); 1H NMR (300 MHz,
D2O, 313 K) (see also Table 5, Supporting Information) δ 3.07
(s, 3 H, NMe), 3.31 (s, 3 H, OMe); 13C NMR (75.5 MHz, D2O,
313 K) δ 184.6, 90.5, 76.2, 73.4, 71.3, 57.5, 46.8, 34.7; FABMS
m/z 237 (30 [M + H]+). Anal. Calcd for C8H16O4N2S: C, 40.66;
H, 6.83; N, 11.86. Found: C, 40.43; H, 6.71; N, 11.53.
1,2,3-Tr i-O-a cetyl-5-d eoxy-5-[3-(2,3,4,6-tetr a -O-a cetyl-
â-D-glu cop yr a n osyl)ca r bod iim id o]-r- a n d -â-D-Xylofu r a -
n ose (44): column chromatography, eluents toluene and then
1:3 f 1:1 EtOAc-petroleum ether; yield 387 mg (82%); Rf (1:1
EtOAc-petroleum ether) 0.57; IR (KBr) νmax 2945, 2145, 1748
cm-1; 1H NMR (500 MHz, CDCl3) (see also Table 4, Supporting
Information) δ 2.15-2.00 (Ac); 13C NMR (125.7 MHz, CDCl3)
δ 170.6-169.1, 138.3, 138.0, 98.7, 92.8, 84.6, 84.5, 80.4, 80.1,
76.3, 75.7, 74.3, 74.1, 73.7, 72.9, 72.7, 68.1, 61.8, 45.9, 45.3,
21.0-20.3; FABMS m/z 653 (100, [M + Na]+). Anal. Calcd for
(2R,3R,4S,5R)-3,4,5-Tr ih yd r oxy-2-m eth oxy-N-(N′-p h en -
ylth ioca r ba m oyl)p ip er id in e (49): column chromatography,
eluent EtOAc f 20:1 EtOAc-EtOH; yield 80 mg (54%); [R]D
+54.0 (c 1.0, MeOH); Rf (45:5:3 EtOAc-EtOH-H2O) 0.47; UV
1
(MeOH) 250 nm (ꢀmM 18.9); H NMR (500 MHz, D2O, 313 K)
(see also Table 5, Supporting Information) δ 7.50-7.30 (m, 5
H, Ph), 3.45 (s, 3 H, OMe); 13C NMR (125.7 MHz, D2O, 313 K)
δ 183.7, 139.5, 129.4, 127.3, 126.9, 88.8, 74.0, 70.8, 69.2, 55.7,
47.9; FABMS m/z 299 (60, [M + H]+). Anal. Calcd for
C13H18O4N3S: C, 52.33; H, 6.08; N, 9.39. Found: C, 52.32; H,
5.98; N, 9.14.
C
26H34O16N2: C, 49.52; H, 5.43; N, 4.44. Found: C, 49.31; H,
5.36; N, 4.35.
Gen er a l P r oced u r e for th e P r ep a r a tion of 5-Deoxy-5-
u r eid o-D-xylofu r a n oses (45-47). To a solution of the cor-
responding carbodiimide 42-44 (0.79 mmol) in 2:1 acetone-
water (15 mL) was added TFA (0.05 mL), and the reaction
mixture was stirred at room temperature for 2.5-5 h (TLC).
After evaporation of the solvents, the residue was purified by
column chromatography eluting first with toluene and then
with the eluent indicated in each case to give the respective
urea 45-47 as an amorphous solid.
1,2,3-Tr i-O-a cetyl-5-d eoxy-5-(3-m eth ylu r eid o)-r- a n d
-â-D-Xylofu r a n ose (45): column chromatography, eluent 3:1
f 4:1 EtOAc-petroleum ether; yield 158 mg (60%); Rf (9:1
CH2Cl2-MeOH) 0.39; IR (KBr) νmax 2938, 1748, 1645, 1566
cm-1; 1H NMR (500 MHz, CDCl3) (see also Table 4, Supporting
Information) δ 5.07-4.95 (NH), 2.70 (d, J Me,N′H ) 5.3 Hz, MeN),
2.09-2.03 (Ac); 13C NMR (125.7 MHz, CDCl3) δ 170.0-168.9,
158.8, 98.9, 92.8, 81.4, 80.3, 77.6, 76.1, 74.6, 74.4, 40.2, 39.4,
26.8, 20.7-20.0; CIMS m/z 333 (55, [M + H]+). Anal. Calcd
for C13H20O8N2: C, 46.948; H, 6.07; N, 8.43. Found: C, 47.13;
H, 6.19; N, 8.51.
(2R,3R,4S,5R)-N-(N′-â-D-Glu copyr an osylth iocar bam oyl)-
3,4,5-tr ih yd r oxy-2-m eth oxyp ip er id in e (50): column chro-
matography, eluent 3:1 MeCN-H2O; yield 169 mg (88%); [R]D
+27.5 (c 1.0, H2O); Rf (6:3:1 MeCN-H2O-NH4OH) 0.41; UV
1
(MeOH) 254 nm (ꢀmM 13.9); H NMR (500 MHz, D2O, 313 K)
(see also Table 5, Supporting Information) δ 3.25 (s, 3 H, OMe);
13C NMR (125.7 MHz, D2O, 313 K) δ 184.5, 88.8, 85.1, 77.6,
76.7, 73.9, 71.8, 70.5, 69.4, 68.7, 60.7, 55.6, 44.9; FABMS m/z
407 (40, [M + Na]+). Anal. Calcd for C13H24O9N2S: C, 40.62;
H, 6.29; N, 7.28. Found: C, 40.55; H, 6.41; N, 7.28.
(2R,3R,4S,5R)-2,3,4,5-Tet r a h yd r oxy-N-(N′-m et h ylca r -
ba m oyl)p ip er id in e (51): column chromatography, eluent
EtOAc f 20:1 EtOAc-EtOH; yield 72 mg (70%); [R]D -2.0 (c
1
1.0, H2O); Rf (4:1 CH2Cl2-MeOH) 0.29; H NMR (500 MHz,
D2O) (see also Table 5, Supporting Information) δ 3.13 (s, 3
H, MeN); 13C NMR (125.5 MHz, D2O) δ 159.2, 77.2, 73.8, 72.1,
69.9, 42.9, 26.5; FABMS m/z 229 (100, [M + Na]+). Anal. Calcd
for C7H13O5N2: C, 40.97; H, 6.39; N, 13.65. Found: C, 40.84;
H, 6.25; N, 13.41.
1,2,3-Tr i-O-a cet yl-5-d eoxy-5-(3-p h en ylu r eid o)-r- a n d
-â-D-Xylofu r a n ose (46): column chromatography, eluent 1:1
EtOAc-petroleum ether; yield 289 mg (93%); Rf (1:1 EtOAc-
petroleum ether) 0.29; IR (KBr) νmax 3339, 3054, 2992, 1750,
(2R,3R,4S,5R)-2,3,4,5-Tet r a h yd r oxy-N-(N′-p h en ylca r -
ba m oyl)p ip er id in e (52): column chromatography, eluent 45:
5:3 EtOAc-EtOH-H2O; yield 87 mg (65%); [R]D -6.0 (c 0.5,
MeOH); Rf (45:5:3 EtOAc-EtOH-H2O) 0.36; 1H NMR (500
MHz, D2O) (see also Table 5, Supporting Information) δ 7.47-
7.31 (m, 5 H, Ph); 13C NMR (75.5 MHz, D2O, 313 K) (see also
Table 2) δ 157.8, 138.1, 129.5, 125.2, 123.3, 77.2, 73.7, 71.9,
69.8, 43.1; FABMS m/z 269 (55, [M + H]+). Anal. Calcd for
1
1657, 1553 cm-1; H NMR (500 MHz, CDCl3) (see also Table
4, Supporting Information) δ 7.29-7.06 (m, Ph, N′H), 5.33 (m,
NH), 2.08-2.04 (Ac); 13C NMR (125.7 MHz, CDCl3) δ 170.2-
169.1, 154.6, 139.1, 129.4, 128.9, 128.8, 98.8, 92.9, 80.0, 79.9,
76.8, 75.8, 74.5, 74.3, 40.0, 39.3, 21.0-20.3; FABMS m/z 417
(100 [M + Na]+). Anal. Calcd for C18H22O8N2: C, 54.82; H, 5.63;
N, 7.10. Found: C, 54.50; H, 5.28; N, 6.89. Samples of the pure
anomers could be obtained in this case by preparative TLC
with the above eluent: R-anomer [R]D +78.0 (c 1.0, CH2Cl2);
â-anomer [R]D +6.0 (c 1.0, CH2Cl2).
C
12H16O5N2: C, 57.73; H, 6.01; N, 10.44. Found: C, 57.84; H,
5.78; N, 10.47.
(2R,3R,4S,5R)-N-(N′-â-D-Glu cop yr a n osylca r b a m oyl)-
2,3,4,5-tetr a h yd r oxyp ip er id in e (53): column chromatogra-
phy, eluent 6:1 f 4:1 MeCN-H2O; yield 106 mg (60%); [R]D
+6.0 (c 1.0, CH2Cl2); Rf (6:3:1 MeCN-H2O-NH4OH) 0.26; 1H
NMR (500 MHz, D2O) (see also Table 5, Supporting Informa-
tion); 13C NMR (125.7 MHz, D2O) δ 158.1, 81.6, 77.3, 76.7, 76.6,
73.3, 71.6, 71.5, 69.4, 69.3, 60.6, 42.4; FABMS m/z 377 (25,
[M + Na]+). Anal. Calcd for C12H22O10N2: C, 40.68; H, 6.26;
N, 7.91. Found: C, 40.63; H, 6.43; N, 7.71.
1,2,3-Tr i-O-a cetyl-5-d eoxy-5-[3-(2,3,4,6-tetr a -O-a cetyl-
â-D-glu cop yr a n osyl)u r eid o]-r a n d -â-D-Xylofu r a n ose (47):
column chromatography, eluent 4:1 CCl4-acetone; yield 480
mg (95%); Rf (2:1 CCl4-acetone) 0.42; IR (KBr) νmax 3370, 2944,
1
1750, 1666, 1555 cm-1; H NMR (500 MHz, CDCl3) (see also
Table 4, Supporting Information) δ 5.51 (d, J N′H,1′ ) 9.5 Hz,
N′HR-anomer), 5.45 (d, J N′H,1′ ) 9.5 Hz, N′Hâ-anomer), 5.13 (dd,
J NH,5b ) 6.7 Hz, J NH,5a ) 6.3 Hz, NHR-anomer), 5.12 (dd, J NH,5a
)
7.2 Hz, J NH,5b ) 4.6 Hz, NHâ-anomer), 2.13-1.99 (Ac); 13C NMR
(125.7 MHz, CDCl3) δ 171.1-169.3, 156.1, 98.9, 92.8, 81.1, 80.2,
77.0, 76.6, 74.6, 74.5, 73.2, 73.1, 72.9, 70.6, 68.9, 68.2, 61.7,
61.5, 40.1, 39.4, 21.1-20.4; FABMS m/z 671 (100, [M + Na]+).
Anal. Calcd for C26H36O17N2: C, 48.15; H, 5.59; N, 4.32.
Found: C, 48.15; H, 5.41; N, 4.25.
Ack n ow led gm en t. M.I.G.-M. thanks the Fundacio´n
Ca´mara for a doctoral fellowship.
Su p p or tin g In for m a tion Ava ila ble: Tables 2-9 contain-
ing 1H and 13C NMR data for all new compounds and the
experimental details for the preparation of the per-O-protected
sugar azides 16 and 36. This material is available free of
charge via the Internet at http://pubs.acs.org.
Gen er a l P r oced u r e for th e P r ep a r a tion of Xylon ojir i-
m ycin -Typ e P ip er id in es (48-53). Conventional NaOMe-
catalyzed deacetylation of (thio)ureas 39-41 and 45-47 (0.5
mmol) in MeOH and purification of the reaction product as
indicated in each case afforded the corresponding N-thiocar-
J O015639F