Synthesis of inositol phosphoglycans containing thiol-terminated spacers for efficient coupling to maleimide functionalized solid phases or proteins

Article abstract of DOI:10.1016/S0040-4020(02)00359-9

The synthesis of inositol phosphoglycans (IPGs), analogous to second messengers of insulin, to provide a small targeted library of compounds is described. These derivatives contain the glucosamine(α1-6)myo-inositol 1,2-cyclic phosphate motif. A thiol-terminated spacer was introduced, for their immobilization, by a radical elongation of an allyl ether with benzyl mercaptane.

Full text of DOI:10.1016/S0040-4020(02)00359-9

TETRAHEDRON
Pergamon
Tetrahedron 58 %2002) 4245±4254
Synthesis of inositol phosphoglycans containing thiol-terminated
spacers for ef®cient coupling to maleimide functionalized solid
phases or proteins
a
Jan Lindberg, Peter Stralfors and Peter Konradsson
b
a,p
Ê
a
È
Department of Chemistry, Linkoping University, SE-581 83 Linkoping, Sweden
Department of Cell Biology, Linkoping University, SE-581 85 Linkoping, Sweden
È
b
È
È
Received 15 January 2002; revised 1 March2002; accepted 28 March2002
AbstractÐThe synthesis of inositol phosphoglycans %IPGs), analogous to second messengers of insulin, to provide a small targeted library
of compounds is described. These derivatives contain the glucosamine%a1±6)myo-inositol 1,2-cyclic phosphate motif. A thiol-terminated
spacer was introduced, for their immobilization, by a radical elongation of an allyl ether with benzyl mercaptane. q 2002 Elsevier Science
Ltd. All rights reserved.
1. Introduction
occur in mammalian cells, although they are not as abundant
as in lower eukaryotes. Mammalian GPIs usually anchor
proteins withspecialized functions. The general structure
of a GPI-anchor is depicted in Fig. 1. It contains an a-linked
non-acetylated glucosamine coupled to phosphatidylinositol
1
Glycosylphosphatidylinositol %GPI)-anchors are anchoring
proteins to the cell membrane and are widely distributed
among eukaryotes. They are common in parasites but also
Figure 1. General structure for GPI-anchors.
Scheme 1. Proposed mechanism for generation of two second messengers.
Keywords: thiol-terminated spacer; inositol phosphoglycans; radical elongation.
Corresponding author. Tel.: 146-13-28-1728; fax: 146-13-28-1399; e-mail: petko@ifm.liu.se
p
0040±4020/02/$ - see front matter q 2002 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020%02)00359-9

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J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
Scheme 2. Coupling of a thiol-functionalized glycan to a solid phase.
at the 6-OH. The glucosamine in all, thus far characterized,
GPI-anchors is substituted at the 4-OH with an identical
ethanolamine±PO4-6Manp%a1±2)Manp%a1±6)Manp%a1±)
core glycan, anchoring the protein at the C-terminus. This
trimannoside is substituted withspecies-speci®c side
chains.
possess insulin-mimetic activity.5 The structure of the
glycan part attached to glucosamine is unknown. However,
insulin mimetic activity of IPGs has been demonstrated in
several studies.4,6 The IPGs were produced from GPI-
anchors by protease digestion and cleavage of the phos-
phodiester bond using bacterial PLC, yielding an inositol
1,2-cyclic phosphate-containing IPG. This has been taken
as an indication of a conserved glycan part, containing
mannose building blocks. Presence of galactose but not
mannose in the active substances has been shown by incor-
poration studies of radiolabeled monosaccharides, indicat-
ing a galactose7,8Ðbut not a mannoseÐcontaining8,9
oligosaccharide attached to the glucosamine. A major
problem in several of these investigations is the poorly
de®ned preparations of IPGs, acquired from natural sources.
Inositol phosphoglycans %IPGs) have been proposed as
second messengers to insulin. They are supposed to be
released from protein-free GPIs in response to insulin, by
the action of a phosphatidylinositol-speci®c phospholipase
C %PLC), together with diacylglycerol which is also acting
as
a
second messenger %Scheme 1).2±4 a-d-Gluco-
samine%1!6)myo-inositol-1,2-cyclic phosphate has been
shown to be the minimum structural motif for the IPGs to
Figure 2. The targeted library of IPGs.

J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
4247
Scheme 3. %a) CHCl3/TFA 10:1; %b) MeOPOCl2, pyridine; %c) Na, NH3 %l); %d) BnSH, AIBN, benzene, 708C; %e) MeOPOCl2, pyridine; %f) Na, NH3 %l).
Therefore a well-de®ned library of synthetic IPGs is neces-
sary in the search for the putative second messenger.
usually give the terminal sul®de with high regioselectivity,
and examples of acetyl or benzyl protected thiols generated
by reaction of allyl ethers with thioacetic acid or benzyl
mercaptane have been reported.10±13 General protecting
group considerations, such as the limited stability of thio-
acetates compared to benzyl sul®des, and ease of deprotec-
tion, makes the benzyl group more attractive as a protecting
group for the thiol. Because of the presence of a sul®de, the
use of catalytic hydrogenation in the deprotection step was
not considered, leaving debenzylation using dissolving
metal reduction in liquid ammonia as the only realistic
alternative.
Furthermore, development of a spacer for coupling of the
IPGs to a solid phase, proteins or various probes will open
up new interesting possibilities, suchas the isolation of
antibodies raised against the IPGs and the identi®cation of
receptors by using photoaf®nity or radiolabeling probes.
The presence of an amino group and a cyclic phosphate in
the target molecules precludes the use of amino- or
carboxylic acid-functionalized spacers. One convenient
way to immobilize the IPGs is therefore to introduce a
thiol-terminated spacer for coupling to a maleimide func-
tionality as depicted in Scheme 2.
The disaccharide 1, together with the three trisaccharides 3,
5, 7 and their spacer containing analogues 2, 4, 6 and 8
%Fig. 2) were chosen as synthetic targets. This small targeted
library of IPGs, is based upon the indications of mannose or
galactose residues in the putative second messenger.
Alkenes are known to react withalkane thiols in radical
reactions to produce dialkyl sul®des. Terminal alkenes
Scheme 4.

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J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
Scheme 5. %a) DMTST, diethyl ether, 15 or 17; %b) CHCl3/TFA 10:1; %c) MeOPOCl2, pyridine; %d) Na, NH3 %l); %e) BnSH, AIBN, benzene, 708C; %f)
MeOPOCl2, pyridine; %g) Na, NH3 %l).
2. Results and discussion
time at elevated temperature was needed for complete
conversion of the allylic double bond to a benzyl sul®de.
Use of smaller excess of the thiol and/or the initiator gave
residual unreacted starting material, which could not be
separated from the product. The resulting compound 12
containing a thiol terminated spacer, protected as a benzyl
sul®de, was obtained in 65% yield. Subsequent phos-
phorylation and deprotection produced the target IPG 2
%Scheme 3).
The camphor acetal of compound 914 was hydrolyzed in
CHCl3/TFA 10:1 to produce compound 10 in 82% yield.
Phosphorylation using N-methylpyridinium dichloro-
phosphate, prepared in situ from methyl dichlorophosphate
in pyridine, produced the cyclic phosphate-containing
derivative 11. Subsequent deprotection and reduction of
the azide using sodium in liquid ammonia gave the target
compound 1. The radical reaction of 10 withbenzyl mer-
captane initiated by 2,20-azobisisobutyronitrile %AIBN) in
benzene at 708C was very slow, and use of a large excess
of both the thiol and AIBN combined with long reaction
The methods developed for the `disaccharide' for introduc-
tion of the spacer and subsequent phospohorylation was
then used to synthesize the IPGs 3±8. The glycosyl donors
Scheme 6. %a) Acrolein dimethyl acetal, DMF; %b) Ac2O, pyridine; %c) NaBH3CN, THF, HCl; %d) Ac2O, pyridine.

J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
4249
Scheme 7. %a) NIS, TfOH, CH2Cl2/diethyl ether 1:1; %b) CHCl3/TFA 10:1; %c) MeOPOCl2, pyridine; %d) MeOH, NH3 %sat.); %e) Na, NH3 %l); %f) BnSH, AIBN,
benzene, 708C; %g) MeOPOCl2, pyridine; %h) MeOH, NH3 %sat.); %i) Na, NH3 %l).
15 and 17 were synthesized by allylation of 14 and 16
%Scheme 4), acquired from mannose or galactose in six
steps.15,16 Subsequent dimethyl%methylthio)sulfonium
tri¯uorometheanesulfonate %DMTST)-promoted glycosyl-
ation of 1814 produced 19 and 20 in 75 and 59% yield,
respectively. Hydrolysis of the camphor acetal gave
compound 21 and 22, which were subjected to the same
transformations as the `disaccharide' 10 to produce the
target IPGs 3±6 %Scheme 5).
The glycosylation of 18 promoted by N-iodosuccinimide
%NIS) and TfOH acid produced trisaccharide 28 in 84%
yield. Hydrolysis of the camphor acetal followed by
phosphorylation or radical elongation and subsequent phos-
phorylation produced 30 and 32. The following deacetyl-
ation had to be performed using mild conditions to preserve
the cyclic phosphate. Methanol saturated with ammonia
accomplished the deacetylation without affecting the cyclic
phosphate. Subsequent debenzylation and reduction of the
azide using sodium in liquid ammonia gave the target
compounds 7 and 8 %Scheme 7).
The b-galactose containing trisaccharides were produced in
the same manner. For construction of the b-galactose-
containing IPGs, an acetylated galactosyl donor was
needed. Other neighbouring participating groups such as
benzoates were ruled out because of the dif®culties asso-
ciated withdeprotection of benzoates in the presence of a
cyclic phosphate. An elegant method to introduce the allyl
ether regioselectively at the 6-OH is to convert the galactose
derivative 25 into the 4,6-O-acrolein acetal derivative 26,
followed by reductive opening of the acetal17 by the same
method as for the more familiar reductive opening of
benzylidene acetals using NaBH3CN. Thus, treatment of
25 withacrolein dimethyl acetal and TsOH, followed by
acetylation produced 26 in 71% yield. Subsequent reaction
3. Experimental
3.1. General methods
Organic phases were dried over %MgSO4), ®ltered, and
concentrated in vacuo at or below 408C. TLC: 0.25 mm
precoated silica-gel plates %MERCK silica-gel 60F254);
detection by spraying the plates 1 M H2SO4 solution
followed by heating at ,2508C. FlashCrhomatograpyh
%FC): Silica gel MERCK 60 %0.040±0.063 mm). Gel
®ltrations were performed on a Pharmacia Sephadex G-15
withNaBH CN and HCl in THF, followed by acetylation
3
column eluted withdeoxygenated %Ar) H O containing 1%
2
gave the donor 27 in 64% yield %Scheme 6).
n-butanol. NMR spectra were recorded on Bruker AC-F 250

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J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
and Varian Mercury 300 spectrometers, temperature 258C.
Chemical shifts are given in ppm relative to TMS in CDCl3
%dˆ0.00) or acetone in D2O %13C: dˆ31.00, 1H: dˆ2.22) as
internal standards; 31P, 85% phosphoric acid %dˆ0.00) was
used as external standard. pHp in D2O is given as an uncor-
rected value calibrated against H2O-buffer solutions. Opti-
cal rotations were recorded at room temperature witha
Perkin±Elmer 241 polarimeter. Melting points were
recorded witha Gallenkamp melting point apparatus.
FAB-mass spectra were recorded on a JEOL SX 102 Mass
Spectrometer using a 3-nitrobenzyl alcohol matrix. IR
spectra were recorded as ®lms on CaF2 crystals %syrups)
or as KBr pellets %solids) on a Perkin±Elmer SPECRUM
1000 FT-IR Spectrometer.
3.1.3. 6-O-[2-Azido-4-O-{3-*benzylthio)propyl}-3,6-di-O-
benzyl-2-deoxy-a-d-glucopyranosyl]- 3,4,5-tri-O-benzyl-
d-myo-inositol *12). A deoxygenated %Ar) mixture of 10
%120 mg, 0.140 mmol), benzylmercaptane %0.53 mL,
4.5 mmol) and AIBN %130 mg, 0.79 mmol) in benzene
%25 mL) was stirred under Ar at 708C. After 11 h, the
mixture was allowed to attain rt and after another 7 h, the
mixture was concentrated. FC %toluene/EtOAc gradient
12:1!2:1) gave 12 %91 mg, 0.093 mmol, 66%) as a color-
less syrup. Rfˆ0.39 %toluene/EtOAc 2:1); [a]Dˆ137 %c
0.52, CHCl3); IR nmax %cm21) 1051, 1453, 2107, 2917;
1
NMR: H %250 MHz, CDCl3), d 1.50±1.70 %2H, m), 2.18±
2.38 %2H, m), 3.00 %1H, dd, Jˆ11.1, 1.8 Hz), 3.15 %1H, dd,
Jˆ11.1, 2.2 Hz), 3.33±4.17 %15H, m), 4.40 %1H, d,
Jˆ11.3 Hz), 4.63±5.01 %8H, m), 5.32 %1H, d, Jˆ3.7 Hz),
7.17±7.33 %30H, m); 13C %62.8 MHz, CDCl3), d 27.9
%CH2), 29.9 %CH2), 36.2 %CH2), 64.3 %CH), 67.4 %CH2),
69.4 %CH), 71.1 %2C, CH, CH2), 72.6 %CH2), 72.8 %CH),
73.3 %CH2), 75.0 %CH2), 75.3 %CH2), 75.9 %CH2), 77.9
%CH), 79.8 %CH), 80.6 %CH), 80.7 %CH), 80.9 %CH), 81.7
%CH), 98.8 %CH), 126.9±128.8 %Ph), 137.7±138.5 %Ph);
HRMS Calcd for C57H63N3O10S: [M1Na]1 1004.4132
Found: [M1Na]1 1004.4181.
3.1.1. 6-O-*4-O-Allyl-2-azido-3,6-di-O-benzyl-2-deoxy-
a-d-glucopyranosyl)-3,4,5-tri-O-benzyl-d-myo-inositol
*10). To 914 %173 mg, 0.174 mmol) in CHCl3 %10 mL) was
added TFA %1 mL). After 16 h, the mixture was diluted with
CHCl3, washed with aqueous NaHCO3 %sat.), dried, ®ltered
and concentrated. FC %toluene/EtOAc 4:1) gave 10 %122 mg,
0.142 mmol, 82%) as a white solid. Rfˆ0.39 %toluene/
EtOAc 2:1); mp 110±1128C %from ether); [a]Dˆ147 %c
1.4, CHCl3); IR nmax %cm21) 1052, 1453, 2107, 2867,
3441; NMR: 1H %250 MHz, CDCl3), d 3.02 %1H, d,
Jˆ9.8 Hz), 3.20 %1H, dd, Jˆ11.1, 2.0 Hz), 3.36 %1H, t,
Jˆ9.3 Hz), 3.43±3.61 %4H, m), 3.83±3.91 %3H, m), 3.95
%1H, t, Jˆ9.5 Hz), 4.01 %1H, t, Jˆ9.5 Hz), 4.09±4.19 %3H,
m), 4.42 %1H, d, Jˆ11.7 Hz), 4.66 %1H, d, Jˆ11.0 Hz), 4.70
%2H, s), 4.78 %1H, d, Jˆ10.6 Hz), 4.81 %1H, d, Jˆ10.6 Hz),
4.88 %1H, d, Jˆ11.3 Hz), 4.92 %1H, d, Jˆ11.0 Hz), 5.01 %1H,
d, Jˆ11.0 Hz), 5.05±5.15 %2H, m), 5.35 %1H, d, Jˆ3.7 Hz),
5.68±5.84 %1H, m), 7.12±7.42 %25H, m); 13C %62.8 MHz,
CDCl3), d 64.2 %CH), 67.4 %CH2), 69.3 %CH), 71.1 %CH),
72.6 %CH2), 72.8 %CH), 73.3 %CH2), 73.5 %CH2), 75.0 %CH2),
75.4 %CH2), 75.8 %CH2), 77.9 %CH), 79.8 %CH), 80.6 %CH),
80.7 %CH), 80.9 %CH), 81.7 %CH), 98.8 %CH), 116.4
%CH2vCH), 127.3±128.5 %Ph), 134.8 %CH2vCH), 137.8
%Ph), 137.9 %Ph), 138.8 %Ph); Anal. Calcd for
C50H55N3O10: C, 70.0; H, 6.5 Found: C, 70.2; H 6.3.
3.1.4. 6-O-*2-Amino-4-O-{3-mercaptopropyl}-2-deoxy-
a-d-glucopyranosyl)-d-myo-inositol 1,2-cyclic phosphate
*2). 12 %17 mg, 0.017 mmol) was phosphorylated and depro-
tected by the same method as for 10 to give 2 %7 mg,
0.015 mmol, 85%) as a white solid. [a]Dˆ130 %c 0.43,
H2O); IR nmax %cm21) 845, 1042, 1099, 1631, 3388;
1
NMR: H %250 MHz, D2O, pHpˆ3.7), d 1.82±1.96 %2H,
m), 2.63 %2H, t, Jˆ7.1 Hz), 3.36±3.46 %3H, m), 3.68 %1H,
t, Jˆ9.9 Hz), 3.72±4.04 %8H, m), 4.56 %1H, ddd, Jˆ20.1,
8.0, 4.8 Hz), 4.71 %1H, t, Jˆ3.8 Hz), 5.50 %1H, d, Jˆ3.7 Hz);
13C %62.8 MHz, D2O, pHpˆ3.7), d 21.1 %CH2), 34.0 %CH2),
55.0 %CH), 60.5 %CH2), 70.2 %CH), 70.3 %CH, d, Jˆ9 Hz).
71.3 %CH), 72.2 %3C, 2£CH, CH2), 77.8 %CH), 78.2 %CH),
80.2 %CH), 81.5 %CH), 96.4 %CH); 31P %121 MHz, D2O,
pHpˆ3.7), d 16.0 %d, Jˆ20.1 Hz); HRMS Calcd for
C15H28NO12PS: [M2H]2 476.0992 Found: [M2H]2
476.0974.
3.1.2.
6-O-*2-Amino-2-deoxy-a-d-glucopyranosyl)-d-
myo-inositol 1,2-cyclic phosphate *1).18 Methyl dichloro-
phosphate %50 mL, 0.50 mmol) was added to pyridine
%1 mL) and the resulting suspension was stirred for
30 min. 10 %38 mg, 0.044 mmol) in pyridine %1 mL) was
added and the mixture stirred for 2 h. NaHCO3 %sat.,
1 mL) was added and the mixture concentrated. The residue
was dissolved in CHCl3 %40 mL) and H2O %10 mL). 1 M
HCl %3 mL) was added and the organic layer separated.
The aqueous layer was extracted with a second portion of
CHCl3 %40 mL) and the combined organic layers were dried
and concentrated to give 11 quantitatively. A solution of 11
in THF %2 mL) was added to NH3 %l) %,20 mL) at 2338C.
To the stirred mixture was added a minimum amount of
sodium for the mixture to turn deep blue. After 1 min,
NH4Cl was added until the color disappeared. The
mixture was concentrated under a stream of argon and the
residue dissolved in H2O %10 mL), washed with ether
%10 mL) and concentrated. Gel ®ltration of the residue
3.1.5. Ethyl 6-O-allyl-2,3,4-tri-O-benzyl-1-thio-a-d-
mannopyranoside *15). 1414 %1.54 g, 3.11 mmol) and
allyl bromide %0.40 mL, 4.7 mmol) dissolved in DMF
%20 mL) was added dropwise over 5 min to 60% NaH
%220 mg, 5.5 mmol) under N2 at 08C. The resulting mixture
was stirred for 2 hat rt. MeOH %1.5 mL) was added and the
mixture was diluted withtoluene, washed withwater, dried
®ltered and concentrated. FC %toluene/EtOAc 20:1) gave 15
%1.62 g, 3.03 mmol 97%) as a colorless syrup. Rfˆ0.42
%toluene/EtOAc 12:1); [a]Dˆ176 %c 1.1, CHCl3); IR nmax
1
%cm21) 1097, 1454, 2869; NMR: H %250 MHz, CDCl3), d
1.23 %3H, t, Jˆ7.3 Hz), 2.46±2.69 %2H, m), 3.66 %1H, dd,
Jˆ11.0, 1.6 Hz), 3.75 %1H, dd, Jˆ11.0, 4.8 Hz), 3.79±3.85
%2H, m), 3.93±4.13 %4H, m), 4.56±4.76 %5H, m), 4.93 %1H,
d, Jˆ11.0 Hz), 5.14 %1H, dd, Jˆ10.2, 1.1 Hz), 5.25 %1H, dd,
Jˆ17.2, 1.5 Hz), 5.39 %1H, s), 5.84±5.99 %1H, m), 7.24±
7.40 %15H, m); 13C %62.8 MHz, CDCl3), d 15.0 %CH3),
25.3 %CH2), 69.2 %CH2), 71.8 %CH), 71.9 %CH2), 72.0
%CH2), 72.3 %CH2), 75.1 %CH), 75.2 %CH2), 76.3 %CH),
80.3 %CH), 81.9 %CH), 116.8 %CH2vCH), 127.6±128.3
%Ph), 134.9 %CH2±CH), 138.1 %Ph), 138.3 %Ph), 138.7 %Ph);
1
afforded 1 %16 mg, 0.040 mmol, 90%) as a white solid. H
and 13C NMR were in accordance withthose previously
published.18

J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
4251
Anal. Calcd for C32H38O5S: C, 71.9; H, 7.2 Found: C, 71.8;
H, 7.1.
ether %10 mL) under argon was added DMTST %233 mg,
0.90 mmol). After 5 h, NEt3 %0.3 mL) was added and after
an additional 15 min, the mixture was ®ltered through Celite
and concentrated. FC %toluene/EtOAc 25:1) gave 20
%172 mg, 0.121 mmol, 59%) as a colorless syrup. Rfˆ0.47
%toluene/EtOAc 12:1); [a]Dˆ172 %c 1.0, CHCl3); IR nmax
%cm21) 1038, 1096, 1454, 2105, 2931; NMR: 1H %250 MHz,
CDCl3), d 0.83 %s, 3H), 0.86 %3H, s), 1.06 %3H, s), 1.15±1.96
%7H, m), 3.31±4.94 %36H, m), 5.11 %1H, dd, Jˆ10.2,
1.8 Hz), 5.19 %1H, dd, Jˆ17.2, 1.5 Hz), 5.58 %1H, d,
Jˆ3.3 Hz), 5.60 %1H, d, Jˆ3.7 Hz), 5.71±5.86 %1H, m),
7.07±7.38 %40H, m); 13C %62.8 MHz, CDCl3), d 9.7 %CH3),
20.4 %CH3), 20.6 %CH3), 27.0 %CH2), 29.8 %CH2), 44.7 %CH2),
45.1 %CH), 47.9 %Cq), 51.6 %Cq), 63.2 %CH), 68.4 %CH2), 68.8
%CH2), 69.5 %CH), 70.3 %CH), 72.1 %CH2), 72.5 %2C, CH2),
73.0 %CH2), 73.4 %CH2), 73.5 %CH2), 73.8 %CH), 74.3 %CH),
74.5 %CH), 74.6 %CH2), 74.7 %CH2), 74.8 %CH2), 75.9 %CH),
76.0 %CH), 76.8 %CH), 78.2 %CH), 79.0 %CH), 80.1 %CH),
80.5 %CH), 80.8 %CH), 95.5 %CH), 98.2 %CH), 116.9
%CH2vCH), 118.0 %OCO), 127.2±128.4 %Ph), 134.5
%CH2vCH), 138.2±138.7 %Ph); Anal. Calcd for
C87H97N3O15: C, 73.3; H, 6.9 Found: C, 73.2; H, 6.8.
3.1.6. Ethyl 6-O-allyl-2,3,4-tri-O-benzyl-1-thio-b-d-
galactopyranoside *17). 1616 %1.23 g, 2.49 mmol) and
allyl bromide %0.32 mL, 3.8 mmol) dissolved in DMF
%20 mL) was added dropwise over 5 min to 60% NaH
%160 mg, 4.0 mmol) under N2 at 08C. The resulting mixture
was stirred for 4 hat rt. MeOH %0.5 mL) was added and the
mixture was diluted withtoluene, washed withwater, dried
®ltered and concentrated. FC %toluene/EtOAc 20:1) gave 17
%1.31 g, 2.45 mmol, 98%) as white needles. Rfˆ0.73
%toluene/EtOAc 4:1); mp 65±668C %from hexane);
[a]Dˆ23.2 %c 1.2, CHCl3); IR nmax %cm21) 1094, 1360,
1
1452, 2920; NMR: H %250 MHz, CDCl3), d 1.30 %3H, t,
Jˆ7.5 Hz), 2.66±2.82 %2H, m), 3.50±3.60 %4H, m), 3.83
%1H, t, Jˆ9.5 Hz), 3.84±3.96 %3H, m), 4.43 %1H, d,
Jˆ9.9 Hz), 4.66 %1H, d, Jˆ11.7 Hz), 4.73 %2H, s), 4.79
%1H, d, Jˆ10.2 Hz), 4.88 %1H, d, Jˆ10.2 Hz), 4.97 %1H, d,
Jˆ11.7 Hz), 5.14±5.27 %2H, m), 5.76±5.92 %1H, m), 7.25±
7.41 %15H, m); 13C %62.8 MHz, CDCl3), d 15.1 %CH3),24.8
%CH2), 68.7 %CH2), 72.3 %CH2), 72.8 %CH2), 73.6 %CH), 74.4
%CH2), 75.8 %CH2), 77.3 %CH), 78.5 %CH), 84.1 %CH), 85.3
%CH), 117.4 %CH2vCH), 127.5±128.5 %Ph), 134.4
%CH2vCH), 138.3 %Ph), 138.4 %Ph), 138.8 %Ph); Anal.
Calcd for C32H38O5S: C, 71.9; H, 7.2 Found: C, 71.8; H 7.3.
3.1.9. 6-O-[4-O-*6-O-Allyl-2,3,4-tri-O-benzyl-a-d-manno-
pyranosyl)-2-azido-3,6-di-O-benzyl-2-deoxy-a-d-gluco-
pyranosyl]-3,4,5-tri-O-benzyl-d-myo-inositol *21). To 19
%242 mg, 0.170 mmol) in CHCl3 %20 mL) was added TFA
%2 mL). After 16 h, the mixture was washed with aqueous
NaHCO3 %sat.), dried, ®ltered and concentrated. FC
%toluene/EtOAc gradient 4:1!2:1) gave 21 %182 mg,
0.141 mmol, 83%) as a colorless syrup. Rfˆ0.46 %toluene/
3.1.7. 6-O-[4-O-*6-O-Allyl-2,3,4-tri-O-benzyl-a-d-manno-
pyranosyl)-2-azido-3,6-di-O-benzyl-2-deoxy-a-d-gluco-
pyranosyl]-3,4,5-tri-O-benzyl-1,2-O-*l-1,7,7-trimethyl-
[2,2,1]bicyclohept-6-ylidene)-d-myo-inositol *19). To a
stirred mixture of 1814 %216 mg, 0.227 mmol), 15 %172 mg,
EtOAc 2:1); [a]Dˆ134 %c 0.8, CHCl3); IR nmax %cm21
)
1
Ê
0.322 mmol) and 4 A molecular sieves in diethyl ether
1046, 1454, 2106, 2867; NMR: H %250 MHz, CDCl3), d
3.34±5.09 %37H, m), 5.18 %1H, dd, Jˆ17.2, 1.5 Hz), 5.24
%1H, d, Jˆ1.8 Hz), 5.53 %1H, d, Jˆ3.3 Hz), 5.75±5.90 %1H,
m), 7.10±7.35 %40H, m); 13C %62.8 MHz, CDCl3), d 64.2
%CH), 68.6 %CH2), 69.1 %CH2), 69.6 %CH), 70.7 %CH), 72.0
%CH2), 72.2 %CH2), 72.3 %CH2), 72.6 %CH2), 72.7 %CH), 72.9
%CH), 73.0 %CH2), 74.3 %CH2), 74.7 %CH), 74.8 %CH2), 75.2
%CH2), 75.8 %CH2), 76.1 %CH), 77.4 %CH), 79.5 %CH), 79.6
%CH), 79.7 %CH), 80.5 %CH), 81.0 %CH), 81.5 %CH), 98.0
%CH, JCHˆ174 Hz), 100.6 %CH, JCHˆ170 Hz), 116.6
%CH2vCH), 126.8±129.0 %Ph), 134.9 %CH2vCH), 137.6±
139.0 %Ph); Anal. Calcd for C77H83N3O15: C, 71.7; H, 6.5
Found: C, 71.8; H 6.6.
%20 mL) under argon was added DMTST %230 mg,
0.89 mmol). After 4 h, NEt3 %0.3 mL) was added and after
an additional 15 min, the mixture was ®ltered through Celite
and concentrated. FC %toluene/EtOAc 25:1) gave 19
%242 mg, 0.170 mmol, 75%) as a colorless syrup. Rfˆ0.47
%toluene/EtOAc 12:1); [a]Dˆ147 %c 1.1, CHCl3); IR nmax
%cm21) 1028, 1108, 1454, 2104, 2933; NMR: 1H %250 MHz,
CDCl3), d 0.83 %3H, s), 0.87 %3H, s), 1.06 %3H, s), 1.15±1.96
%7H, m), 3.35 %1H, dd, Jˆ9.7, 3.5 Hz), 3.38±4.77 %33H, m),
4.88 %1H, d, Jˆ11.3 Hz), 4.91 %1H, d, Jˆ11.0 Hz), 5.06±
5.26 %3H, m), 5.62 %1H, d, Jˆ3.3 Hz), 5.77±5.92 %1H, m),
7.02±7.36 %40H, m); 13C %62.8 MHz, CDCl3), d 9.7 %CH3),
20.4 %CH3), 20.6 %CH3), 27.0 %CH2), 29.8 %CH2), 44.7 %CH2),
45.1 %CH), 47.9 %Cq), 51.6 %Cq), 63.1 %CH), 68.4 %CH2), 69.2
%CH2), 70.1 %CH), 72.1 %CH2), 72.2 %CH2), 72.3 %CH2), 72.5
%CH2), 72.9 %CH), 73.0 %CH2), 73.8 %CH), 73.9 %CH2), 74.7
%CH2), 74.8 %CH), 74.9 %CH2), 75.0 %CH2), 76.0 %CH), 76.3
%CH), 76.8 %CH), 77.6 %CH), 78.0 %CH), 79.6 %CH), 79.8
%CH), 80.5 %CH), 80.9 %CH), 95.4 %CH, JCHˆ174 Hz),
100.9 %CH, JCHˆ174 Hz), 116.6 %CH2vCH), 118.1
%OCO), 126.9±128.5 %Ph), 134.9 %CH2vCH), 137.8±
138.6 %Ph); Anal. Calcd for C87H97N3O15: C, 73.3; H, 6.9
Found: C, 73.5; H 6.9.
3.1.10.
6-O-[4-O-*6-O-Allyl-2,3,4-tri-O-benzyl-a-d-
galactopyranosyl)-2-azido-3,6-di-O-benzyl-2-deoxy-a-d-
glucopyranosyl]-3,4,5-tri-O-benzyl-d-myo-inositol *22).
To 20 %154 mg, 0.108 mmol) in CHCl3 %15 mL) was
added TFA %1.5 mL). After 16 h, the mixture was washed
withaqueous NaHCO %sat.), dried, ®ltered and concen-
3
trated. FC %toluene/EtOAc 2:1) gave 22 %112 mg,
0.0868 mmol, 80%) as a colorless syrup. Rfˆ0.46 %toluene/
EtOAc 2:1); [a]Dˆ143 %c 1.1, CHCl3); IR nmax %cm21
)
1
1046, 1454, 2108, 2870; NMR: H %250 MHz, CDCl3), d
3.24±5.32 %38H, m), 5.31 %1H, d, Jˆ3.3 Hz), 5.54 %1H, d,
Jˆ3.7 Hz), 5.67±5.80 %1H, m), 7.18±7.36 %40H, m); 13C
%62.8 MHz, CDCl3), d 64.9 %CH), 68.3 %CH2), 68.8 %CH2),
69.2 %CH), 69.8 %CH), 71.4 %CH), 72.1 %CH2), 72.4 %CH),
72.5 %2C, CH2), 73.0 %CH2), 73.7 %CH), 73.9 %CH2), 74.2
%CH2), 74.5 %CH), 74.6 %CH2), 74.8 %CH2), 75.7 %CH), 75.8
%CH2), 79.1 %CH), 79.7 %CH), 81.0 %CH), 81.1 %CH), 81.7
3.1.8. 6-O-[4-O-*6-O-Allyl-2,3,4-tri-O-benzyl-a-d-galac-
topyranosyl)-2-azido-3,6-di-O-benzyl-2-deoxy-a-d-
glucopyranosyl]-3,4,5-tri-O-benzyl-1,2-O-*l-1,7,7-tri-
methyl[2,2,1]bicyclohept-6-ylidene)-d-myo-inositol *20).
To a stirred mixture of 18 %195 mg, 0.205 mmol), 17
Ê
%200 mg, 0.374 mmol) and 4 A molecular sieves in diethyl

4252
J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
%CH), 82.0 %CH), 98.0 %CH), 99.1 %CH), 117.1 %CH2vCH),
125.3±129.0 %Ph), 134.5 %CH2vCH), 137.8±138.7 %Ph);
Anal. Calcd for C77H83N3O15: C, 71.7; H, 6.5 Found: C,
71.6; H 6.6.
3.1.14. 6-O-[4-O-*6-O-{3-*Benzylthio)propyl}-2,3,4-tri-
O-benzyl-a-d-galactopyranosyl)-2-azido-3,6-di-O-benzyl-
2-deoxy-a-d-glucopyranosyl]-3,4,5-tri-O-benzyl-d-myo-
inositol *24). 22 %116 mg, 0.090 mmol) was reacted with
benzylmercaptane by the same method as for 10 to give
24 %80 mg, 0.057 mmol, 63%) as a colorless syrup.
Rfˆ0.46 %toluene/EtOAc 2:1); [a]Dˆ137 %c 1.1, CHCl3);
IR nmax %cm21) 1046, 1453, 1495, 2107, 2870, 2917; NMR:
1H %250 MHz, CDCl3), d 1.63±1.68 %2H, m), 2.30±2.39
%2H, m), 3.14±5.00 %38H, m), 5.31 %1H, d, Jˆ3.7 Hz),
5.54 %1H, d, Jˆ3.7 Hz), 7.17±7.34 %45H, m); 13C
%62.8 MHz, CDCl3), d 28.0 %CH2), 29.3 %CH2), 36.3
%CH2), 65.0 %CH), 68.7 %CH2), 68.9 %CH2), 69.2 %CH),
69.6 %CH2), 69.7 %CH), 71.5 %CH), 72.3 %CH), 72.5 %CH2),
72.6 %CH2), 73.0 %CH2), 73.5 %CH), 74.0 %CH2), 74.3 %CH2),
74.5 %2C, CH, CH2), 74.8 %CH2), 75.7 %CH) 75.9 %CH2), 79.1
%CH), 79.8 %CH), 81.1 %CH), 81.2 %CH), 81.7 %CH), 82.3
%CH), 98.0 %CH), 99.2 %CH), 126.9±129.0 %Ph), 137.8±
138.5 %Ph); HRMS Calcd for C84H91N3O15S: [M1Na]1
1436.6069 Found: [M1Na]1 1436.6064.
3.1.11. 6-O-[4-O-*a-d-Mannopyranosyl)-2-amino-2-deoxy-
a-d-glucopyranosyl]-d-myo-inositol 1,2-cyclic phosphate
*3).19,20 21 %38 mg, 0.029 mmol) was phosphorylated and
deprotected by the same method as for 10 to give 3
%14 mg, 0.025 mmol, 84%) as a white solid. [a]Dˆ175 %c
0.6, H2O), Lit.19 [a]Dˆ157 %c 1.0, H2O); IR nmax %cm21
)
1
845, 1042, 1099, 1221, 1633, 3375; NMR: H %250 MHz,
D2O, pHpˆ5.4), d 3.37±3.44 %2H, m), 3.65±4.21 %14H, m),
4.56 %1H, ddd, Jˆ20.1, 8.0, 4.8 Hz), 4.71 %1H, t, Jˆ4.0 Hz),
5.28 %1H, d, Jˆ1.5 Hz), 5.51 %1H, d, Jˆ3.3 Hz); 13C
%62.8 MHz, D2O, pHpˆ5.4), d 55.0 %CH), 60.7 %CH2),
61.6 %CH2), 67.3 %CH), 70.2 %CH, d, Jˆ11.1 Hz), 70.6
%CH), 71.0 %CH), 71.1 %CH), 71.4 %CH), 71.8 %CH), 72.2
%CH), 74.5 %CH), 76.5 %CH), 77.8 %CH), 80.2 %CH), 81.6
%CH), 96.4 %CH), 102.3 %CH); 31P %121 MHz, D2O,
pHpˆ5.4), d 16.0 %d, Jˆ20.1 Hz); HRMS Calcd for
C18H32NO17P: [M2H]2 564.1330 Found: [M2H]2
564.1367.
3.1.15. 6-O-[4-O-*6-O-{3-Mercaptopropyl}-a-d-manno-
pyranosyl)-2-amino-2-deoxy-a-d-glucopyranosyl]-d-myo-
inositol 1,2-cyclic phosphate *4). 23 %37 mg, 0.026 mmol)
was phosphorylated and deprotected by the same method as
for 10 to give 4 %15 mg, 0.023 mmol, 90%) as a white solid.
[a]Dˆ166 %c 0.7, H2O); IR nmax %cm21) 845, 1039, 1102,
3.1.12.
deoxy-a-d-glucopyranosyl]-d-myo-inositol
6-O-[4-O-*a-d-Galactopyranosyl)-2-amino-2-
1,2-cyclic
phosphate *5). 22 %31 mg, 0.024 mmol) was phosphoryl-
ated and deprotected by the same method as for 10 to give
5 %12 mg, 0.021 mmol, 88%) as a white solid. [a]Dˆ1106
%c 0.8, H2O); IR nmax %cm21) 844, 1033, 1095, 1222, 1634,
1
1222, 1635, 3225; NMR: H %250 MHz, D2O, pHpˆ4.7), d
1.85±1.97 %2H, m), 2.62 %2H, t, Jˆ7.1 Hz), 3.37±3.44 %2H,
m), 3.61±4.14 %16H, m), 4.56 %1H, ddd, Jˆ20.1, 8.0,
4.8 Hz), 4.71 %1H, t, Jˆ4.0 Hz), 5.26 %1H, s) 5.51 %1H, d,
Jˆ3.7 Hz); 13C %62.8 MHz, D2O, pHpˆ4.7), d 21.2 %CH2),
33.4 %CH2), 55.0 %CH), 60.8 %CH2), 67.4 %CH), 70.2 %CH, d,
Jˆ9 Hz), 70.3 %2C, CH2), 70.6 %CH), 70.9 %CH), 71.0 %CH),
71.3 %CH), 71.9 %CH), 72.2 %CH), 73.3 %CH), 76.6 %CH),
77.8 %CH), 80.2 %CH), 81.5 %CH), 96.3 %CH), 102.3 %CH);
31P %121 MHz, D2O, pHpˆ4.7), d 16.2 %d, Jˆ20.1 Hz);
HRMS Calcd for C21H38NO17PS: [M2H]2 638.1520
Found: [M2H]2 638.1493.
1
3405; NMR: H %250 MHz, D2O, pHpˆ5.6), d 3.38±3.45
%2H, m), 3.65±4.25 %14H, m), 4.57 %1H, ddd, Jˆ20.1, 8.0,
4.8 Hz), 4.71 %1H, t, Jˆ4.0 Hz), 5.45 %1H, d, Jˆ2.6 Hz),
5.52 %1H, d, Jˆ3.7 Hz); 13C %62.8 MHz, D2O, pHpˆ5.6),
d 54.8 %CH), 60.7 %CH2), 61.9 %CH2), 69.3 %CH), 69.9
%CH), 70.0 %CH), 70.3 %CH2, d, Jˆ9.2 Hz), 70.7 %CH),
71.4 %CH), 71.6 %CH), 72.2 %CH), 72.5 %CH), 76.6 %CH),
77.8 %CH), 80.3 %CH), 81.4 %CH), 96.3 %CH), 100.4 %CH);
31P %121 MHz, D2O, pHpˆ5.6), d 16.0 %d, Jˆ20.1 Hz);
HRMS Calcd for C18H32NO17P: [M2H]2 564.1330
Found: [M2H]2 564.1266.
3.1.16. 6-O-[4-O-*6-O-{3-Mercaptopropyl}-a-d-galacto-
pyranosyl)-2-amino-2-deoxy-a-d-glucopyranosyl]-d-myo-
inositol 1,2-cyclic phosphate *6). 24 %39 mg, 0.028 mmol)
was phosphorylated and deprotected by the same method as
for 10 to give 6 %15 mg, 0.024 mmol, 85%) as a white solid.
[a]Dˆ176 %c 0.6, H2O); IR nmax %cm21) 845, 1036, 1102,
3.1.13. 6-O-[4-O-*6-O-{3-*Benzylthio)propyl}-2,3,4-tri-
O-benzyl-a-d-mannopyranosyl)-2-azido-3,6-di-O-benzyl-
2-deoxy-a-d-glucopyranosyl]-3,4,5-tri-O-benzyl-d-myo-
inositol *23). 21 %88 mg, 0.068 mmol) was reacted with
benzylmercaptane by the same method as for 10 to give
23 %52 mg, 0.037 mmol, 54%) as a colorless syrup.
Rfˆ0.51 %toluene/EtOAc 2:1); [a]Dˆ139 %c 0.86,
CHCl3); IR nmax %cm21) 1048, 1453, 2106, 2867; NMR:
1H %250 MHz, CDCl3), d 1.60±1.80 %2H, m), 2.40 %2H, t,
Jˆ7.1 Hz), 3.29±4.96 %38H, m), 5.24 %1H, d, Jˆ1.8 Hz),
5.48 %1H, d, Jˆ3.6 Hz), 7.10±7.40 %45H, m); 13C
%62.8 MHz, CDCl3), d 28.1 %CH2), 29.4 %CH2), 36.2
%CH2), 64.4 %CH), 68.6 %CH2), 69.6 %CH), 69.7 %CH2),
70.0 %CH2), 70.9 %CH), 72.1 %CH2), 72.3 %CH2), 72.7
%CH2), 72.8 %CH), 72.9 %CH), 73.2 %CH2), 74.5 %CH2),
74.7 %CH), 74.9 %CH2), 75.1 %CH2), 75.8 %CH2), 76.1
%CH), 77.2 %CH), 79.6 %CH), 79.8 %CH), 80.1 %CH), 80.6
%CH), 81.0 %CH), 81.6 %CH), 98.3 %CH), 100.5 %CH),
126.9±129.0 %Ph), 137.6±138.9 %Ph); HRMS Calcd for
C84H91NO15S: [M1Na]1 1436.6069 Found: [M1Na]1
1436.6083.
1
1224, 1634, 3225; NMR: H %250 MHz, D2O, pHpˆ4.1), d
1.84±1.94 %2H, m), 2.61 %2H, t, Jˆ7.1 Hz), 3.38±3.45 %2H,
m), 3.60±4.24 %16H, m), 4.57 %1H, ddd, Jˆ20.1, 8.0,
4.8 Hz), 4.71 %1H, t, Jˆ4.0 Hz), 5.43 %1H, d, Jˆ2.2 Hz),
5.52 %1H, d, Jˆ3.7 Hz); 13C %62.8 MHz, D2O, pHpˆ4.1),
d 21.1 %CH2), 33.3 %CH2), 54.8 %CH), 60.8 %CH2), 69.2
%CH), 69.9 %CH), 70.2 %CH), 70.2 %CH, d, Jˆ9 Hz), 70.3
%CH2), 70.6 %CH), 70.7 %CH2), 70.9 %CH), 71.3 %CH), 71.7
%CH), 72.1 %CH), 76.9 %CH), 77.8 %CH), 80.2 %CH), 81.5
%CH), 96.3 %CH), 100.5 %CH); 31P %121 MHz, D2O,
pHpˆ4.1), d 16.2 %d, Jˆ20.1 Hz); HRMS Calcd for
C21H38NO17PS: [M2H]2 638.1520 Found: [M2H]2
638.1613.
3.1.17. Ethyl 2,3,-di-O-acetyl-4,6-O-prop-2-enylidene-1-
thio-b-d-galactopyranoside *26). A solution of 25
%1.50 g, 6.70 mmol), acrolein dimethylacetal %1.50 mL,

J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
4253
12.7 mmol) and TsOH´H2O %100 mg) in DMF %5 mL), was
stirred at 508C. After 1.5 h, pyridine %10 mL) was added and
the mixture allowed to attain rt. Acetic anhydride %5 mL)
was added and after an additional 3 h, the mixture was
diluted withCH 2Cl2 and washed with H2O, 1 M HCl,
NaHCO3 and H2O, dried ®ltered and concentrated. FC
%toluene/EtOAc 4:1) gave 26 %1.64 g, 4.73 mmol, 71%) as
colorless needles. Rfˆ0.30 %toluene/EtOAc 4:1); mp 139±
1408C %from ethanol); [a]Dˆ25.0 %c 1.4, CHCl3); IR nmax
%cm21) 826, 927, 955, 991, 1057, 1093, 1171, 1223, 1243,
0.183 mmol, 78%) as a colorless syrup. Rfˆ0.57 %toluene/
)
EtOAc 4:1); [a]Dˆ136 %c 0.8, CHCl3); IR nmax %cm21
1057, 1220, 1753, 2107, 2936; NMR: 1H %250 MHz,
CDCl3), d 0.88 %6H, s), 1.06 %3H, s), 1.16±2.0 %7H, m),
1.72 %3H, s), 1.95 %3H, s), 2.11 %3H, s), 3.19±3.28 %2H,
m), 3.32 %1H, dd, Jˆ10.2, 3.7 Hz), 3.44±4.10 %13H, m),
4.30 %1H, m), 4.37 %1H, d, Jˆ12.1 Hz), 4.50 %1H, d,
Jˆ8.1 Hz), 4.54 %1H, d, Jˆ11.3 Hz), 4.57±4.80 %8H, m),
5.04±5.20 %4H, m), 5.34 %1H, d, Jˆ2.9 Hz), 5.54 %1H, d,
Jˆ3.7 Hz), 5.66±5.81 %1H, m), 7.17±7.46 %25H, m); 13C
%62.8 MHz, CDCl3), d 9.9 %CH3), 20.4 %CH3), 20.5 %CH3),
20.6 %CH3), 20.7 %CH3), 21.4 %CH3), 27.0 %CH2), 29.9 %CH2),
44.4 %CH2), 45.2 %CH), 47.9 %Cq), 51.6 %Cq), 63.0 %CH), 66.9
%CH2), 67,3 %CH2), 67.6 %CH), 69.8 %CH), 70.4 %CH), 71.4
%CH), 72.0 %CH), 72.3 %CH2), 72.7 %CH2), 73.7 %2C, CH,
CH2), 73.8 %CH2), 74.1 %CH2), 74.8 %CH2), 75.8 %CH),
76.4 %2C, CH), 77.4 %CH), 78.6 %CH), 80.4 %CH), 80.5
%CH), 96.0 %CH), 100.0 %CH), 117.2 %CH2vCH),
117.9%OCO), 127.4±129.0 %Ph), 134.3 %CH2vCH), 137.6
%Ph), 137.8 %Ph), 138.2 %Ph), 138.3 %Ph), 138.4 %Ph), 138.7
%Ph), 169.1 %CH3CO), 169.9 %CH3CO), 170.0 %CH3CO);
Anal. Calcd for C72H85N3O18: C, 67.5; H, 6.7 Found: C,
67.3; H 6.9.
1
1376, 1742, 2985; NMR: H %250 MHz, CDCl3), d 1.28
%3H, t, Jˆ7.5 Hz), 2.07 %3H, s), 2.09 %3H, s), 2.63±2.91
%2H, m), 3.48 %1H, d, Jˆ1.1 Hz), 3.87 %1H, dd, Jˆ12.6,
1.6 Hz), 4.15±4.31 %2H, m), 4.43 %1H, d, Jˆ9.9 Hz),
4.91±4.96 %2H, m) 5.33 %1H, d, Jˆ10.6 Hz), 5.43 %1H, t,
Jˆ10.1 Hz), 5.48 %1H, d, Jˆ17.5 Hz), 5.91 %1H, ddd,
Jˆ17.5, 10.6, 4.7 Hz); 13C %62.8 MHz, CDCl3), d 14.8
%CH3), 20.9 %2C, CH3), 23.1 %CH2), 66.7 %CH), 68.7
%CH2), 69.8 %CH), 73.0 %CH), 73.2 %CH), 82.9 %CH), 100.5
%OCO), 119.3 %CH2vCH), 133.9 %CH2vCH), 169.5
%CH3CO), 170.7 %CH3CO); Anal. Calcd for C15H22O7S: C,
52.0; H, 6.4 Found: C, 52.0; H 6.5.
3.1.18. Ethyl 6-O-allyl-2,3,4-tri-O-acetyl-1-thio-b-d-
galactopyranoside *27). To a stirred mixture of 26
%268 mg, 0.774 mmol) and NaBH3CN %495 mg, 7.9 mmol)
in THF %10 mL) ether saturated with HCl was added drop-
wise until all starting material was consumed. The mixture
3.1.20. 6-O-[4-O-*6-O-Allyl-2,3,4-tri-O-acetyl-b-d-galac-
topyranosyl)-2-azido-3,6-di-O-benzyl-2-deoxy-a-d-gluco-
pyranosyl]-3,4,5-tri-O-benzyl-d-myo-inositol *29). To 28
%160 mg, 0.125 mmol) in CHCl3 %15 mL) was added TFA
%1.5 mL). After 16 h, the mixture was diluted with CHCl3,
washed with aqueous NaHCO3 %sat.), dried, ®ltered and
concentrated. FC %toluene/EtOAc 2:1) gave 29 %124 mg,
0.108 mmol, 86%) as a colorless syrup. Rfˆ0.28 %toluene/
1056, 1220, 1750, 2116, 2871; NMR: 1H %250 MHz,
CDCl3), d 1.77 %3H, s), 1.94 %3H, s), 2.09 %3H, s), 3.03±
4.08 %17H, m), 4.23 %1H, br), 4.29 %1H, d, Jˆ8.0 Hz), 4.55
%1H, d, Jˆ11.8 Hz), 4.64±4.73 %6H, m), 4.89 %1H, d,
Jˆ10.6 Hz), 4.98±5.21 %m, 6H) 5.79 %1H, d, Jˆ2.9 Hz),
5.65±5.80 %1H, m) 7.14±7.44 %25H, m); 13C %62.8 MHz,
CDCl3), d 20.6 %CH3), 20.7 %2C, CH3), 64.2 %CH), 66.6
%CH2), 66.8 %CH2), 67.4 %CH), 69.0 %CH), 69.7 %CH), 71.1
%CH), 71.2 %CH), 71.9 %CH), 72.2 %CH2), 72.3 %CH), 72.5
%CH2), 73.5 %CH2), 73.6 %CH2), 75.2 %CH2), 75.7 %CH), 75.9
%CH2), 78.8 %CH), 79.8 %CH), 80.6 %CH), 81.6 %CH), 83.1
%CH), 99.6 %CH), 99.8 %CH), 117.3 %CH2vCH), 126.5 %Ph),
127.3±128.6 %Ph), 134.3 %CH2vCH), 137.3 %Ph), 137.8
%Ph), 138.3 %2C, Ph), 138.8 %Ph), 169.2 %CH3CO), 169.9
%CH3CO), 170.0 %CH3CO); Anal. Calcd for C62H71N3O18:
C, 65.0; H, 6.2 Found: C, 64.8; H 6.2.
was neutralized withNEt %2 mL) and passed through a
3
short column of silica eluted with toluene/EtOAc 1:1. The
crude mixture was concentrated and dissolved in pyridine
%5 mL). Ac2O %2 mL) was added to the stirred mixture. After
3 h, the mixture was concentrated. Puri®cation by FC
%petroleum ether 65±75/EtOAc 6:1) gave 27 %193 mg,
0.494 mmol, 64%) as a colorless syrup. Rfˆ0.34 %toluene/
EtOAc 2:1); [a]Dˆ111 %c 1.4, CHCl3); IR nmax %cm21
)
EtOAc 4:1); [a]Dˆ217 %c 1.0, CHCl3); IR nmax %cm21
)
1
1222, 1749; NMR: H %250 MHz, CDCl3), d 1.28 %3H, t,
Jˆ7.5 Hz), 1.98 %3H, s), 2.07 %3H, s), 2.15 %3H, s), 2.63±
2.84 %2H, m), 3.4 %1H, dd, Jˆ9.7, 6.4 Hz), 3.56 %1H, dd,
Jˆ9.8, 6.2 Hz), 3.86 %1H, dt, Jˆ6.2, 0.7 Hz), 3.90 %1H,
dddd, Jˆ12.8, 5.8, 1.3, 1.3 Hz) 3.99 %1H, dddd, Jˆ12.8,
6.5, 1.3, 1.3 Hz), 4.50 %1H, d, Jˆ9.9 Hz), 5.06 %1H, dd,
Jˆ9.9, 3.3 Hz), 5.15±5.28 %3H, m), 5.49 %1H, d,
Jˆ2.6 Hz), 5.75±5.91 %1H, m); 13C %62.8 MHz, CDCl3), d
14.9 %CH3), 20.6 %CH3), 20.7 %CH3), 20.8 %CH3), 24.4 %CH2),
67.5 %CH), 67.8 %CH2), 68.0 %CH), 72.2 %CH), 72.4 %CH2),
76.0 %CH), 84.0 %CH), 117.6 %CH2vCH), 134.2
%CH2vCH), 169.6 %CH3CO), 170.1 %CH3CO), 170.2
%CH3CO); Anal. Calcd for C17H26O8S: C, 52.3; H, 6.7
Found: C, 52.4; H 6.9.
3.1.21.
deoxy-a-d-glucopyranosyl]-d-myo-inositol
6-O-[4-O-*b-d-Galactopyranosyl)-2-amino-2-
1,2-cyclic
3.1.19. 6-O-[4-O-*6-O-Allyl-2,3,4-tri-O-acetyl-b-d-galac-
topyranosyl)-2-azido-3,6-di-O-benzyl-2-deoxy-a-d-gluco-
pyranosyl]-3,4,5-tri-O-benzyl-1,2-O-*l-1,7,7-trimethyl-
[2,2,1]bicyclohept-6-ylidene)-d-myo-inositol *28). To a
stirred mixture of 27 %160 mg, 0.410 mmol), 18 %224 mg,
phosphate *7). 29 %25 mg, 0.020 mmol) was phosphoryl-
ated by the same method as for 10 to give 30. The product
was dissolved in MeOH/NH3 %sat., 2 mL). After 6 h, the
mixture was concentrated and dissolved in THF %2 mL).
NH3 %l) %,20 mL) was condensed into the ¯ask and a
minimum amount of sodium for the mixture to turn deep
blue was added to the stirred mixture at 2338C. After 1 min,
NH4Cl was added until the color disappeared. The mixture
was concentrated under a stream of argon and the residue
was dissolved in H2O %10 mL), washed with ether %10 mL)
and concentrated. Gel ®ltration of the residue on a
Ê
0.235 mmol) and 4 A powdered molecular sieves in
CH2Cl2/ether 1:1 %5 mL) at 08C under argon was added
NIS %98 mg, 0.44 mmol) and TfOH %0.20 mL, 1% in
CH2Cl2). After 20 min, the mixture was diluted with
CH2Cl2, ®ltered through celite, washed with NaHCO3
%sat.) and 1 M NaS2O3, dried, ®ltered and concentrated.
FC %toluene/EtOAc gradient 12:1!6:1) gave 28 %234 mg,

4254
J. Lindberg et al. / Tetrahedron 58 (2002) 4245±4254
Pharmacia Sephadex G-15 column eluted with H2O contain-
ing 1% n-butanol afforded 7 %11 mg, 0.020 mmol, quant.) as
a white solid. [a]Dˆ149 %c 0.42, H2O); IR nmax %cm21) 848,
%CH2), 33.2 %CH2), 54.8 %CH), 60.1 %CH2), 69.0 %CH),
69.5 %CH), 69.8 %CH2), 70.2 %CH, d, Jˆ9.2 Hz), 70.4
%CH2), 71.3 %CH), 71.6 %CH), 71.7 %CH), 72.2 %CH), 73.2
%CH), 73.9 %CH), 77.8 %CH), 78.7 %CH), 80.2 %CH), 81.5
%CH), 96.2 %CH), 103.8 %CH); 31P %121 MHz, D2O,
pHpˆ4.7), d 16.2 %d, Jˆ20.1 Hz); HRMS Calcd for
C21H38NO17PS: [M2H]2 638.1520 Found: [M2H]2
638.1614.
1
1038, 1095, 1223, 1634, 3188; NMR: H %250 MHz, D2O,
pHpˆ5.2), d 3.38±4.16 %16H, m), 4.48 %1H, d, Jˆ7.7 Hz)
4.56 %1H, ddd, Jˆ20.1, 8.0, 4.8 Hz), 4.71 %1H, t, Jˆ4.0 Hz),
5.51 %1H, d, Jˆ3.7 Hz); 13C %62.8 MHz, D2O, pHpˆ5.2), d
54.7 %CH), 60.1 %CH2), 61.9 %CH2), 69.0 %CH), 69.3 %CH),
70.2 %CH, d, Jˆ10.1 Hz), 71.3 %CH), 71.7 %2C, CH), 72.1
%CH), 73.3 %CH), 76.2 %CH), 77.8 %CH), 78.4 %CH), 80.2
%CH), 81.5 %CH), 96.2 %CH), 103.8 %CH); 31P %121 MHz,
D2O, pHpˆ5.2), d 16.0 %d, Jˆ20.1 Hz); HRMS Calcd for
C18H32NO17P: [M2H]2 564.1330 Found: [M2H]2
564.1282.
References
1. Anderson, R. G. Sem. Immunol. 1994, 6, 89±95.
2. Jones, D. R.; Varela-Nieto, I. Mol. Med. 1999, 5, 505±514.
Ê
3. Stralfors, P. Nature 1988, 335, 554±556.
Ê
4. Stralfors, P. BioEssays 1997, 19, 327±335.
3.1.22. 6-O-[4-O-*6-O-{3-*Benzylthio)propyl}-2,3,4-tri-
O-acetyl-b-d-galactopyranosyl)-2-azido-3,6-di-O-benzyl-
2-deoxy-a-d-glucopyranosyl]-3,4,5-tri-O-benzyl-d-myo-
inositol *31). 29 %69 mg, 0.060 mmol) was reacted with
benzylmercaptane by the same method as for 10 to give
31 %48 mg, 0.038 mmol, 63%) as a colorless syrup.
Rfˆ0.28 %toluene/EtOAc 2:1); [a]Dˆ111 %c 1.1, CHCl3);
5. Misek, D. E.; Saltiel, A. R. J. Biol. Chem. 1992, 267, 16266±
16273.
Ê
6. Alemany, S.; Mato, J. M.; Stralfors, P. Nature 1987, 330, 77±
79.
7. Parpal, S.; Gustavsson, J.; Stralfors, P. J. Cell Biol. 1995, 131,
Ê
125±135.
8. Suzuki, S.; Sugawara, K.; Satoh, Y.; Toyota, T. J. Biol. Chem.
1991, 266, 8115±8121.
1
IR nmax %cm21) 1055, 1221, 1751, 2114, 2917; NMR: H
%250 MHz, CDCl3), d 1.54 %2H, m), 1.77 %3H, s), 1.94 %3H,
s), 2.10 %3H, s), 2.34±2.40 %2H, m), 3.02±5.10 %32H, m),
5.19 %1H, d, Jˆ3.7 Hz), 5.28 %1H, d, Jˆ3.3 Hz), 7.14±7.43
%30H, m); 13C %62.8 MHz, CDCl3), d 20.6 %3C, CH3), 27.9
%CH2), 29.0 %CH2), 36.2 %CH2), 64.3 %CH), 66.7 %CH2), 67.4
%CH2), 67.5 %CH), 69.0 %CH), 69.8 %2C, CH, CH2), 71.1
%CH), 71.3 %CH), 71.8 %CH), 72.3 %CH), 72.6 %CH2), 73.5
%CH2), 73.7 %CH2), 75.2 %CH2), 75.8 %CH), 75.9 %CH2), 78.8
%CH), 79.8 %CH), 80.6 %CH), 81.7 %CH), 83.2 %CH), 99.7
%CH), 99.8 %CH), 125.3 %Ph), 126.5±129.1 %Ph), 137.4±
138.9 %Ph), 169.3 %CH3CO), 169.9 %2C, CH3CO); HRMS
Calcd for C69H79N3O18S: [M1Na]1 1292.4977 Found:
[M1Na]1 1292.4998.
9. Mato, J. M.; Kelly, K. L.; Abler, A.; Jarett, L. J. Biol. Chem.
1987, 262, 2131±2137.
10. Matsuoka, K.; Oka, H.; Koyama, T.; Esumi, Y.; Terunuma, D.
Tetrahedron Lett. 2001, 42, 3327±3330.
11. Roy, R.; Baek, M.-G.; Rittenhouse-Olson, K. J. Am. Chem.
Soc. 2001, 123, 1809±1816.
12. Matsuoka, K.; Kurosawa, H.; Esumi, Y.; Terunuma, D.;
Kuzuhara, H. Carbohydr. Res. 2000, 329, 765±772.
13. Plourde, R.; Johnson, L.; Longo, R. Synth. Lett. 2001, 3, 439±
441.
È
14. Lindberg, J.; Ohberg, L.; Garegg, P. J.; Konradsson, P.
Tetrahedron 2002, 58, 1387±1398.
15. Ottosson, H. Carbohydr. Res. 1990, 197, 101±107.
16. Ruda, K.; Lindberg, J.; Garegg, P. J.; Oscarson, S.;
Konradsson, P. J. Am. Chem. Soc. 2000, 122, 11067±11072.
17. Garegg, P. J.; Hultberg, H.; Oscarson, S. J. Chem. Soc. Perkin
Trans. 1 1982, 2395±2397.
3.1.23. 6-O-[4-O-*6-O-{3-Mercaptopropyl}-b-d-galacto-
pyranosyl)-2-amino-2-deoxy-a-d-glucopyranosyl]-d-myo-
inositol 1,2-cyclic phosphate *8). 31 %31 mg, 0.024 mmol)
was phosphorylated and deprotected by the same method as
for 29 to give 8 %14 mg, 0.022 mmol, 90%) as a white solid.
[a]Dˆ140 %c 0.83, H2O); IR nmax %cm21) 845, 1039, 1097,
18. Plourde, R.; d'Alarcao, M.; Saltiel, A. R. J. Org. Chem. 1992,
57, 2606±2610.
1
1221, 1634, 3152; NMR: H %250 MHz, D2O, pHpˆ4.7), d
19. Dietrich, H.; Espinosa, J. F.; Chiara, J. L.; Jimenez-Barbero,
J.; Leon, Y.; Varela-Nieto, I.; Mato, J. M.; Cano, F. H.; Foces-
1.84±1.94 %2H, m), 2.62 %2H, t, Jˆ7.1 Hz), 3.38±3.45 %2H,
m), 3.51±4.16 %16H, m), 4.48 %1H, d, Jˆ7.7 Hz), 4.57 %1H,
ddd, Jˆ20.1, 8.0, 4.7 Hz), 4.71 %1H, t, Jˆ4.0 Hz), 5.52 %1H,
d, Jˆ3.7 Hz); 13C %62.8 MHz, D2O, pHpˆ4.7), d 21.1
Â
Foces, C.; Martõn-Lomas, M. Chem. Eur. J. 1999, 5, 320±336.
20. Jaworek, C. H.; Calias, P.; Iacobucci, S.; d'Alarcao, M.
Tetrahedron Lett. 1999, 40, 667±670.