Synthesis of a C-glycoside analogue of β-D-galactosylthreonine

Article abstract of DOI:10.1021/jo026758d

A C-linked analogue of β-D-galactosylthreonine has been prepared from 2,3,4,6-tetra-O-benzyl-D-galactopyranolactone (1) in 14 steps. Three stereogenic centers were created during the synthesis, with the anomeric center of the C-glycoside being generated f

Full text of DOI:10.1021/jo026758d

Previous studies have thus focused on naturally occurring
glycosylated amino acids where the amino acid moiety
contains only one stereogenic center.12 However, glyco-
proteins also carry a variety of saccharides linked to
amino acids which have two stereogenic centers. For
instance, in mucins which are found on epithelial cells,
threonine carries R-D-N-acetylgalactosamine residues to
which larger saccharide structures are often attached.13
Nuclear pore proteins, transcriptional factors, and cy-
toskeletal proteins have â-D-N-acetylglucosamine moi-
eties attached to threonine.14 Hydroxylysine, found in the
collagens, is often glycosylated with â-D-galactosyl moi-
eties.15 As part of a project directed toward C-glycosidic
analogues of these two amino acids we now report the
synthesis of a C-linked analogue of â-D-galactosylated
threonine (cf. 13, Scheme 2). Galactosylated threonine
residues are found in the cuticle collagen of the deep-sea
hydrothermal vent worm Riftia pachyptila, and were
recently demonstrated to contribute to the relatively high
thermal stability of this protein.16
Synthesis of a C-linked analogue of â-D-galactosylated
threonine requires stereocontrol at three centers, namely,
the anomeric carbon of the C-glycoside and the two
stereogenic centers of the threonine moiety. In the
present work the configuration at the anomeric center
was established first by synthesis of the â-linked C-
glycoside 2 (Scheme 1). De novo synthesis of the threo-
nine part was then carried out by building on the
propanol moiety of 2. The two key steps, in which the
threonine stereogenic centers were established, both
relied on alkylation of enolates derived from N-acyl-
oxazolidinones, as developed by Evans.17
Syn th esis of a C-Glycosid e An a logu e of
â-D-Ga la ctosylth r eon in e
Tomas Gustafsson, Maria Saxin, and J an Kihlberg*
Organic Chemistry, Department of Chemistry,
Umeå University, SE-901 87 Umeå, Sweden
jan.kihlberg@chem.umu.se
Received November 24, 2002
Abstr a ct: A C-linked analogue of â-D-galactosylthreonine
has been prepared from 2,3,4,6-tetra-O-benzyl-D-galacto-
pyranolactone (1) in 14 steps. Three stereogenic centers were
created during the synthesis, with the anomeric center of
the C-glycoside being generated first by addition of a
Grignard reagent to 1 and subsequent reduction of the
intermediate hemiacetal with triethylsilane. The two ste-
reogenic centers in the threonine moiety were both estab-
lished by alkylation of Evans’ chiral N-acyloxazolidinone
enolates.
Most proteins are post-translationally modified by
glycosylation, with O-glycosylation being found on serine,
threonine, hydroxylysine, and tyrosine while N-glycosy-
lation occurs on asparagine.1,2 Glycosylation influences
properties of proteins such as solubility, stability toward
proteases, as well as conformation and folding. It can also
affect the biological functions of proteins in events of
molecular recognition, e.g., cell-to-cell communication and
adhesion of bacteria or viruses to cell-surface proteins.
In recent years, there has been an increasing interest in
the synthesis of carbon-linked analogues of glycosylated
amino acids, in which a methylene group has replaced
the O- or N-glycosidic linkage.3 Such C-glycosides mimic
the native glycosides in many ways such as conformation
and size, but differ in other aspects such as electrostatic
and chemical properties.4 In contrast to O-glycosides,
C-glycosides are not degraded under acidic conditions or
by glycosidases, nor do they undergo base-catalyzed
â-elimination, which constitutes a problem for glycosides
of serine and threonine. This makes C-glycosides inter-
esting in efforts to modulate biological properties of
glycopeptides and glycoproteins and suggests applications
in drug discovery.
The synthetic route started by addition of homoallyl-
magnesium bromide to lactone 1,18 followed by reduction
of the intermediate hemiacetal with triethylsilane and
boron trifluoride etherate (Scheme 1). Ozonolyzis of the
resulting alkene and decomposition of the ozonide with
NaBH4 gave the â-linked C-glycoside 219 (88% from 1),
which provided carboxylic acid 3 in excellent yield (99%)
upon J ones oxidation of the propanol moiety. Carboxim-
ide 4 was then prepared (70% yield) by conversion of 3
into a mixed pivaloyl anhydride, which was immediately
reacted with Evans chiral auxiliary, (R)-4-benzyloxazo-
lidinone. Alkylation of 4 established the stereochemistry
(10) Dondoni, A.; Marra, A.; Massi, A. J . Org. Chem. 1999, 64, 933-
944.
(11) Dondoni, A.; Mariotti, G.; Marra, A. Tetrahedron Lett. 2000,
41, 3483-3487.
Several syntheses of C-linked analogues of glycosylated
amino acids found in Nature have been reported, includ-
ing different C-glycosides of serine3,5-8 and asparagine.3,9-11
(12) C-Linked analogues of glycosylated amino acids, where the
amino acid part contains two stereogenic centers, but which are not
found in nature have been prepared. Cf. Westermann, B.; Walter, A.;
Flo¨rke, U.; Altenbach, H.-J . Org. Lett. 2001, 3, 1375-1378.
(13) Carraway, K. L.; Hull, S. R. Glycobiol. 1991, 1, 131-138.
(14) Zachara, N. E.; Hart, G. W. Chem. Rev. 2002, 102, 431-438.
(15) Kivirikko, K. I.; Myllyla¨, R. In Collagen in health and disease;
Weiss, J . B., J ayson, M. I. V., Eds.; Churchill Livingstone: Edinburgh,
1982; pp 101-120.
(1) Lis, H.; Sharon, N. Eur. J . Biochem. 1993, 218, 1-27.
(2) Dwek, R. A. Chem. Rev. 1996, 96, 683-720.
(3) Dondoni, A.; Marra, A. Chem. Rev. 2000, 100, 4395-4421.
(4) Levy, D. E.; Tang, C. The chemistry of C-glycosides; Elsevier
Science Ltd: Oxford, 1995; Vol. 13.
(5) Bertozzi, C. R.; Cook, D. G.; Kobertz, W. R.; Gonzalez-Scarano,
F.; Bednarski, M. D. J . Am. Chem. Soc. 1992, 114, 10639-10641.
(6) Lay, L.; Meldal, M.; Nicotra, F.; Panza, L.; Russo, G. Chem.
Commun. 1997, 1469-1470.
(16) Bann, J . G.; Peyton, D. H.; Ba¨chinger, H. P. FEBS Lett. 2000,
473, 237-240.
(7) Urban, D.; Skrydstrup, T.; Beau, J .-M. Chem. Commun. 1998,
955-956.
(17) Evans, D. A.; Ennis, M. D.; Mathre, D. J . J . Am. Chem. Soc.
1982, 104, 1737-1739.
(8) Dondoni, A.; Mariotti, G.; Marra, A.; Massi, A. Synthesis 2001,
2129-2137.
(18) Dondoni, A.; Scherrmann, M.-C. J . Org. Chem. 1994, 59, 6404-
6412.
(9) Campbell, A. D.; Paterson, D. E.; Raynham, T. M.; Taylor, R. J .
K. Chem. Commun. 1999, 1599-1600.
(19) Wellner, E.; Gustafsson, T.; Ba¨cklund, J .; Holmdahl, R.; Kihl-
berg, J . ChemBioChem 2000, 1, 272-280.
10.1021/jo026758d CCC: $25.00 © 2003 American Chemical Society
Published on Web 02/25/2003
2506
J . Org. Chem. 2003, 68, 2506-2509

SCHEME 1a
SCHEME 2a
a
Reagents and conditions: (a) three steps, see ref 19 (88%); (b)
a
CrO3/H2SO4, acetone, rt (99%); (c) (i) Et3N, Me3CCOCl, THF, -78
°C; (ii) (R)-4-benzyloxazolidin-2-one, BuLi, THF, -78 °C (70%); (d)
LDA, allyl iodide, THF, -40 to -20 °C (60%); (e) LiEt3BH, THF,
rt (94%); (f) I2, PPh3, imidazole, toluene, rt (90%); (g) LiEt3BH,
THF, rt (96%).
Reagents and conditions: (a) (i) O3, CH2Cl2/MeOH, -78 °C,
(ii) NaBH4, CH2Cl2/MeOH, -78 °C to rt, (iii) CrO3/H2SO4, acetone,
rt (93%); (b) (i) Et3N, Me3CCOCl, THF, -78 °C, (ii) (R)-4-
benzyloxazolidin-2-one, BuLi, THF, -78 °C (79%); (c) (i) DIPEA,
Bu2BOTf, CH2Cl2, -78 °C to rt, (ii) NBS, CH2Cl2, -78 °C (57%);
(d) tetramethylguanidinium azide, CH2Cl2, rt (97%); (e) LiOH,
THF/H2O, rt (97%).
at the â-carbon of the threonine moiety, and was ac-
complished by treatment with LDA and either of allyl
bromide or allyl iodide. This gave 5 in 60% yield and
excellent diastereoselectivety (>99:1), both of which were
unaffected by the choice of electrophile. However, the
alkylation proceeded considerably faster with the more
reactive allyl iodide. The modest yield in this step is most
likely explained by the observation that carboximide 4
decomposed during alkylation, even at -78 °C. At-
tempted reduction of oxazolidinone 5 with LiAlH4 was
surprisingly sluggish and gave, apart from 6, the inter-
mediate aldehyde which was almost inert, even to an
excess of LiAlH4. This was unexpected since the allylated
product obtained from the (S)-benzyloxazolidinone ana-
logue of 4 was easily reduced with LiAlH4. Reduction of
5 was instead performed with LiEt3BH, which provided
a clean reduction to give alcohol 6 (94%). Deoxygenation
of this alcohol, and the corresponding iodide 7, was
attempted with several methods. Tosylation of 6 and
reduction with either LiAlH4 or LiEt3BH20 was unsuc-
cessful, as was reduction of iodide 7 using radical
conditions (Bu3SnH, AIBN) or hydrides (LiAlH4, or
NaBH4 under phase-transfer conditions21). However,
LiEt3BH was successfully employed to convert iodide 7
to the methyl analogue 8 in near quantitative yield.
In compound 8, the desired stereochemistry has been
established for the C-glycosidic linkage, and for the
methyl group which corresponds to the â-carbon of the
threonine moiety. To complete the synthesis of the target
C-linked analogue of â-D-galactosyl threonine it remains
to convert the alkene part of 8 into an L-amino acid
moiety. This was accomplished by first performing an
oxidative cleavage of the alkene moiety using ozone,
followed by reduction with NaBH4 and J ones oxidation
of the resulting alcohol, to provide carboxylic acid 9 in
93% yield (Scheme 2). Compound 9 was then transformed
into oxazolidinone 10 (79%) using the same condidtions
as for 4. Direct azidation of 10 to give 12 (enolization by
KHMDS, then treatment with trisyl azide and acetic
acid) proved to be difficult and a two-step procedure was
therefore used instead. First a boron enolate was pre-
pared from 10 and employed in an electrophilic bromi-
nation with N-bromosuccinimide to give 11. Then nu-
cleophilic displacement of the bromine atom in 11 with
tetramethylguanidinium azide22 gave azido oxazolidinone
12 in 54% yield over these two steps. The stereoselectivity
in the bromination step was somewhat disappointing
since a 93:7 mixture of the two diastereomers was
formed. However, the minor diastereomer could easily
be removed by silica gel flash chromatography, either at
the bromo (11) or the azido stage (12). Surprisingly, both
11 and 12 decomposed somewhat on the column during
purification (silica gel or alumina), but a higher yield was
obtained if 11 was purified before conversion into 12.
Finally, the synthesis was completed by hydrolysis of
oxazolidinone 12 using aqueous LiOH which provided
azido acid 13 in 97% yield. Building block 13 is ready
for use in solid-phase synthesis since it is known that
R-azido acids can be activated and coupled to peptides
in excellent yields without racemization.23 The azido
(20) Krishnamurthy, S.; Brown, H. C. J . Org. Chem. 1976, 41, 3064-
3066.
(21) Rolla, F. J . Org. Chem. 1981, 46, 3909-3911.
(22) Papa, A. J . J . Org. Chem. 1966, 31, 1426-1430.
J . Org. Chem, Vol. 68, No. 6, 2003 2507

group can then be reduced on the solid support,23,24
followed by further elongation of the peptide chain and
deprotection of the benzyl ethers during or after cleavage
of the C-linked glycopeptide from the solid phase.
combined organic phases were dried over Na2SO4, filtered, and
concentrated. Purification by flash column chromatography
(heptane/ethyl acetate 1:1) gave 3 (1.43 g, 99%) as a colorless
oil: [R]D ) -6.3 (c 1.0, CHCl3); HR-MS (FAB) calcd for C37H40
-
NaO7 619.2672 [M + Na]+, found 619.2680.
Epimerization could potentially have occurred at C-R
of the threonine moiety, either during nucleophilic sub-
stitution of 11 or during base-catalyzed hydrolysis of 12
to give 13. To investigate the stereochemical purity of
13 a mixture of 13 and epi-13 (i.e., 13 that had been
epimerized at the C-R position) was synthesized. This was
done by treatment of bromide 11 with tetrabutylammo-
nium bromide in refluxing THF to form a ∼1:1 mixture
of 11 and epi-11, as determined by 1H NMR spectroscopy.
This mixture was treated with tetramethylguanidinium
azide, and hydrolysis of the oxazolidinone, as described
for 12, then provided a ∼1:1 mixture of 13 and epi-13.
Analytical reversed-phase HPLC displayed two peaks for
this mixture, one of which had the same retention time
(R )-4-Be n zyl-3-[3-(2,3,4,6-t e t r a -O-b e n zyl-â-D-ga la ct o-
p yr a n osyl)p r op ion yl]oxa zolid in -2-on e (4). Triethylamine
(1.26 mL, 9.05 mmol) was added to a solution of carboxylic acid
3 (4.91 g, 8.23 mmol) in THF (80 mL) cooled to -78 °C. The
solution was stirred for 5 min, and pivaloyl chloride (1.11 mL,
9.05 mmol) was added. After 15 min, a mixture of (R)-4-
benzyloxazolidin-2-one (1.60 g, 9.05 mmol) and BuLi (3.62 mL,
2.5 M in hexanes, 9.05 mmol) in THF (30 mL) was transferred
to this solution via cannula. The mixture was stirred for 30 min
at -78 °C, and then the reaction was quenched by addition of
NH4Cl (aq, satd). After extraction twice with CH2Cl2, the
combined organic phases were dried over Na2SO4, filtered and
concentrated. Purification by flash column chromatography
(heptane/ethyl acetate 3:1) gave 4 (4.37 g, 70%) as a colorless
oil: [R]D ) -27.8 (c 1.0, CHCl3); HR-MS (FAB) calcd for C47H49
-
NNaO8 778.3356 [M + Na]+, found 778.3354.
1
as that of 13. In addition H NMR spectroscopy revealed
(R)-4-Ben zyl-3-[2-(R)(2,3,4,6-tetr a -O-ben zyl-â-D-ga la cto-
p yr a n osylm eth yl)p en t-4-en oyl]oxa zolid in -2-on e (5). LDA
(6.18 mL, 1.0 M in hexanes, 6.18 mmol) was added to a solution
of oxazolidinone 4 (4.25 g, 5.62 mmol) in THF (50 mL) cooled to
-78 °C. After 30 min, allyl iodide (1.80 mL, 19.7 mmol) was
added, and the temperature was raised to -20 °C during 2 h.
The solution was poured into NH4Cl (aq, satd), the phases were
separated, and the organic phase was washed twice with brine.
The combined aqueous phases were extracted twice with CH2-
Cl2, after which the combined organic phases were dried over
Na2SO4, filtered, and concentrated. Purification by flash column
chromatography (heptane/ethyl acetate 3:1) gave 5 (5.44 g, 62%)
as an amorphous solid: [R]D ) -27.3 (c 1.0, CHCl3); HR-MS
two sets of signals for the mixture, one of which matched
the spectrum of 13. By using HPLC and 1H NMR
spectroscopy it could thus be conclusively determined
that epimerization had not occurred during the azide
displacement of 11, or in the hydrolysis to give 13.
In conclusion, a synthesis of the C-linked analogue of
â-D-galactosyl threonine, 13, has been accomplished from
1 in 12% total yield over fourteen steps. The synthesis
involved creation of three stereogenic centers, with the
anomeric center of the C-glycoside being generated first.
The configuration of the two stereogenic centers in the
threonine moiety of 13 was established in key steps
relying on alkylation of enolates derived from N-acylox-
azolidinones. We anticipate that building blocks such as
13 will be useful in studies of the unexpectedly high
thermal stability of cuticle collagen from the deep-sea
hydrothermal vent worm Riftia pachyptila.
(FAB) calcd for
818.3676.
C
50H53NNaO8 818.3669 [M + Na]+, found
(R)-2-(2,3,4,6-Tetr a -O-ben zyl-â-D-ga la ctop yr a n osylm eth -
yl)p en t-4-en -1-ol (6). LiEt3BH (712 µL, 1 M in THF, 0.712
mmol) was added to a solution of oxazolidinone 5 (189 mg, 0.237
mmol) in THF (1.5 mL) previously cooled to 0 °C. After 20 min,
EtOH (1 mL), H2O (0.5 mL), NaOH (0.5 mL, 5 M), and H2O2
(0.3 mL, 30% aq) was added, and stirring was continued for 5
min. The mixture was poured onto NaCl (aq, satd) and was
extracted three times with CH2Cl2. The combined organic phases
were dried over Na2SO4, filtered, and concentrated. Purification
by flash column chromatography (heptane/ethyl acetate 3:1) gave
6 (139 mg, 94%) as an amorphous solid: [R]D ) -1.7 (c 1.0,
CHCl3); HR-MS (FAB) calcd for C40H46NaO6 645.3192 [M +
Na]+, found 645.3200.
Exp er im en ta l Section
Gen er a l P r oced u r es. All reactions were carried out under
an inert atmosphere with dry solvents under anhydrous condi-
tions, unless otherwise stated. CH2Cl2 was distilled from calcium
hydride, whereas THF and toluene were distilled from potassium
benzophenone and sodium, respectively. Methanol was dried
over 3 Å molecular sieves. TLC was performed on silica gel 60
F254 (Merck) with detection by UV light and staining with a
solution of ethanolic phosphomolybdic acid. Flash column chro-
matography (eluents given in brackets) was performed on silica
(R)-4-(2,3,4,6-Tet r a -O-b en zyl-â-D-ga la ct op yr a n osylm e-
t h yl)-5-iod op en t -1-en e (7). I2 (1.64 g, 6.47 mmol), triphen-
ylphosphine (2.14 g, 8.17 mmol), and imidazole (0.62 g, 9.19
mmol) were added to a solution of alcohol 6 (2.12 g, 3.40 mmol)
in toluene (70 mL). After 20 min, H2O2 (1 mL, 30% aq) was
added, and stirring was continued for 1 min. The resulting
mixture was poured into NaHSO3 (10% aq) and extracted twice
with toluene. The combined organic phases were washed with
NaHCO3 (aq, satd.) and brine, then dried over Na2SO4, filtered,
and concentrated. Flash column chromatography (heptane/ethyl
acetate 4:1) gave iodide 7 (2.25 g, 90%) as a slightly yellow oil:
[R]D ) -8.6 (c 1.0, CHCl3); HR-MS (FAB) calcd for C40H45INaO5
755.2209 [M + Na]+, found 755.2208.
(R)-4-(2,3,4,6-Tetr a -O-ben zyl-â-D-ga la ctop yr a n osylm eth -
yl)p en t-1-en e (8). LiEt3BH (8.35 mL, 1 M in THF, 8.35 mmol)
was added to a solution of 6 (2.04 g, 2.78 mmol) in THF (65 mL)
and stirred for 20 min. Then EtOH (15 mL, 95%), H2O (7 mL),
NaOH (5 mL, 5 M aq), and H2O2 (3 mL, 30% aq) were added.
After being stirred for 10 min, the mixture was poured into brine
and extracted twice with ethyl acetate. The combined organic
phases were dried over Na2SO4, filtered, and concentrated to
afford 8 (1.63 g, 96%) as a colorless oil. Alkene 6 was sufficiently
pure to be used directly in the next step. A small amount of 8
was purified by flash column chromatography (heptane/ethyl
1
gel (Matrex, 60 Å, 35-70 µm, Grace Amicon). H and 13C NMR
spectra were recorded at 400 and 100 MHz, respectively, for
solutions in CDCl3 [residual CHCl3 (δH 7.26 ppm) or CDCl3 (δC
77.0 ppm), as internal standard] at 298 K. First-order chemical
shifts and coupling constants were obtained from one-dimen-
sional spectra; carbon and proton resonances were assigned from
COSY and HETCOR experiments. Resonances for aromatic
hydrogen and carbon atoms are not reported.
3-(2,3,4,6-Tetr a -O-ben zyl-â-D-ga la ctop yr a n osyl)p r op ion -
ic Acid (3). J ones’ reagent (aq 1 M CrO3, 4.4 M H2SO4) was
added to a solution of alcohol 2 (1.42 g, 2.44 mmol) dissolved in
acetone (50 mL) at 0 °C. The mixture was allowed to attain rt
and was then stirred for an additional 90 min. i-PrOH (4 mL)
was added, and the solution was brought to pH 4 with NaHCO3
(aq, satd). It was extracted twice with diethyl ether, and the
(23) Tornoe, C. W.; Davis, P.; Porreca, F.; Meldal, M. J . Peptide Sci.
2000, 6, 594-602.
(24) Brickmann, K.; Yuan, Z.; Sethson, I.; Somfai, P.; Kihlberg, J .
Chem. Eur. J . 1999, 5, 2241-2253.
2508 J . Org. Chem., Vol. 68, No. 6, 2003

acetate 2:1): [R]D ) -19.2 (c 1.0, CHCl3); HR-MS (FAB) calcd
for C39H44NaO7 629.3243 [M + Na]+, found 629.3241.
2 h, NaHSO3 (5% aq) was added, and the solution was extracted
three times with ethyl acetate. The combined organic phases
were washed with NaHSO3 (5% aq) and brine, dried over Na2-
SO4, filtered, and concentrated. Purification by flash column
chromatography (heptane/ethyl acetate 3:1) gave 11 (151 mg,
57%) as a slightly yellow oil: [R]D ) -29.2 (c 1.0, CHCl3); HR-
MS (FAB) calcd for C49H52BrNNaO8 884.2774 [M + Na]+, found
884.2790.
(R)-4-Ben zyl-3-[(2S,3R)-a zid o-3-(2,3,4,6-tetr a -O-ben zyl-â-
D-ga la ctop yr a n osylm eth yl)bu tyr yl]oxa zolid in -2-on e (12).
A solution of bromide 11 (38 mg, 0.044 mmol) and tetrameth-
ylguanidinium azide (21 mg, 0.132 mmol) in CH2Cl2 (0.5 mL)
was stirred for 24 h. Concentration followed by purification by
flash column chromatography (heptane/ethyl acetate 3:1) gave
azide 12 (35 mg, 97%) as an amorphous solid: [R]D ) -12.2 (c
1.0, CHCl3); HR-MS (FAB) calcd for C49H52N4NaO8 847.3683 [M
+ Na]+, found 847.3695.
(2S,3S)-Azid o-3-(2,3,4,6-tetr a -O-ben zyl-â-D-ga la ctop yr a -
n osylm eth yl)bu tyr ic Acid (13). LiOH (606 µL, 0.10 M aq, 61
µmol) was added to a solution of carboximide 12 (25.1 mg, 30
µmol) in THF (1.5 mL) cooled to 0 °C. After 5 min, the solution
was neutralized with Amberlite IR120, filtered and concentrated.
Flash column chromatography (heptane/ethyl acetate 3:1) gave
carboxylic acid 13 (19.2 mg, 97%) as an amorphous solid: [R]D
) -14.2 (c 1.0, CHCl3); HR-MS (FAB) calcd for C39H43N3NaO8
688.2999 [M + Na]+, found 688.3017.
(S)-3-(2,3,4,6-Tetr a -O-ben zyl-â-D-ga la ctop yr a n osylm eth -
yl)bu tyr ic Acid (9). Alkene 8 (1.60 g, 2.64 mmol) was dissolved
in CH2Cl2 (35 mL) and MeOH (30 mL), and the solution was
cooled to -78 °C. O3 (g) was bubbled through the solution until
it turned slightly blue, and then O2 (g) was passed through until
the solution became colorless. NaBH4 (0.30 g, 7.91 mmol) was
added, and the mixture was allowed to attain rt and then stirred
for an additional 30 min. The reaction was quenched by addition
of HCl (5% aq), after which the mixture was poured into brine
and extracted twice with CH2Cl2. The combined organic phases
were dried over Na2SO4, filtered and concentrated. The residue
was dissolved in acetone (40 mL), J ones’ reagent (aq 1 M CrO3,
4.4 M H2SO4) was added at 0 °C, and the mixture was stirred
at rt for 5 min. i-PrOH (4 mL) was added, and the solution was
brought to pH 4 with NaHCO3 (aq, satd). It was then extracted
three times with diethyl ether. The combined organic phases
were dried over Na2SO4, filtered, and concentrated, after which
the crude product was purified by flash column chromatography
(heptane/ethyl acetate 2:1) to afford 9 (1.52 g, 93%) as a colorless
oil: [R]D ) -5.8 (c 1.0, CHCl3); HR-MS (FAB) calcd for C39H44
-
NaO7 647.2985 [M + Na]+, found 647.2985.
(R)-4-Ben zyl-3-[3-(S)-(2,3,4,6-tetr a -O-ben zyl-â-D-ga la cto-
p yr a n osylm eth yl)bu tyr yl]oxa zolid in -2-on e (10). Carboxylic
acid 9 (1.45 g, 2.32 mmol) was reacted with (R)-4-benzyloxazo-
lidin-2-one (452 mg, 2.55 mmol) as described in the preparation
of 4. After workup, purification by flash column chromatography
(heptane/ethyl acetate 3:1) gave 10 (1.43 g, 79%) as an amor-
phous solid: [R]D ) -22.0 (c 4.0, CHCl3); HR-MS (FAB) calcd
for C49H53NNaO8 806.3669 [M + Na]+, found 806.3664.
Ack n ow led gm en t. We thank Dr. Magnus J ohans-
son at the Department of Environmental Chemistry at
Umeå University for performing MS analysis. This work
was funded by grants from the Swedish Research
Council and the Go¨ran Gustafsson Foundation for
Research in Natural Sciences and Medicine.
(R)-4-Ben zyl-3-[(2R,3S)-br om o-(2,3,4,6-tetr a -O-ben zyl-â-
D-ga la ctop yr a n osylm eth yl)bu tyr yl]oxa zolid in -2-on e (11).
Diisopropylethylamine (64 µL, 0.367 mmol) and dibutylboron-
triflate (337 µL, 1 M in CH2Cl2, 0.337 mmol) were added to a
solution of carboximide 10 (240 mg, 0.306 mmol) in CH2Cl2 (1
mL) cooled to -78 °C. After 15 min, the solution was warmed
to 0 °C and stirred for an additional 1 h. The solution was cooled
to -78 °C and transferred to a suspension of NBS (109 mg, 0.612
mmol) in CH2Cl2 (1 mL), which was precooled to -78 °C. After
Su p p or tin g In for m a tion Ava ila ble: Copies of 1H and 13C
NMR spectra for compounds 3-13, as well as listed 1H and
13C NMR data for these compounds. This material is available
free of charge via the Internet at http://pubs.acs.org.
J O026758D
J . Org. Chem, Vol. 68, No. 6, 2003 2509