Lipid A-type pyrancarboxylic acid derivatives, their synthesis and their biological activities

Article abstract of DOI:10.1016/S0040-4020(00)00684-0

Synthesis of lipid A-type pyrancarboxylic acid derivatives, which have a carboxylic acid group in the anomeric position of the reducing sugar part of the disaccharide instead of the phosphoric acid group in lipid A, is described. We investigated the influence of the substituents in the 2'- and 6'-position of the molecules synthesized on their activities toward human monoblastic U937 cells. It was revealed that a series of compounds, possessing an acetamido group in the 2'-position showed strong LPS-antagonistic activity. (C) 2000 Elsevier Science Ltd.

Full text of DOI:10.1016/S0040-4020(00)00684-0

TETRAHEDRON
Pergamon
Tetrahedron 56 (2000) 7691±7703
Lipid A-Type Pyrancarboxylic Acid Derivatives, their Synthesis
and their Biological Activities
c
c
Takashi Mochizuki,a Yuji Iwano,b Masao Shiozaki,a, Shin-ichi Kurakata, Saori Kanai and
*
Masahiro Nishijimad
aExploratory Chemistry Research Laboratories, Sankyo Co. Ltd., Hiromachi 1-2-58, Shinagawa-ku, Tokyo 140-8710, Japan
bChemTech Labo, Inc., Hiromachi 1-2-58, Shinagawa-ku, Tokyo 140-8710, Japan
cBiological Research Laboratories, Sankyo Co. Ltd., Hiromachi 1-2-58, Shinagawa-ku, Tokyo 140-8710, Japan
dDepartment of Biochemistry and Cell Biology, National Institute of Infectious Diseases, Toyama 1-23-1, Shinjuku-ku,
Tokyo 162-8640, Japan
Received 18 May 2000; accepted 2 August 2000
AbstractÐSynthesis of lipid A-type pyrancarboxylic acid derivatives, which have a carboxylic acid group in the anomeric position of the
reducing sugar part of the disaccharide instead of the phosphoric acid group in lipid A, is described. We investigated the in¯uence of the
substituents in the 20- and 60-position of the molecules synthesized on their activities toward human monoblastic U937 cells. It was revealed
that a series of compounds, possessing an acetamido group in the 20-position showed strong LPS-antagonistic activity. q 2000 Elsevier
Science Ltd. All rights reserved.
Introduction
whether the molecules show LPS-agonistic or -antagonistic
activities.10
Lipopolysaccharides (LPS)1 cover the outer surface
membrane of Gram-negative bacteria. LPS are known to
stimulate the immune system of the host cells, resulting in
many pathophysiological events such as fever, depression of
blood pressure, platelet aggregation, shock, and organ
failure leading to bacterial sepsis.2 Most of the biological
activities of LPS reside in a relatively small portion of the
molecule, that is, the terminal disaccharide phospholipid
subunit known as lipid A (1, Fig. 1),1,3,4 which is a hydro-
phobic anchor substance that links an essentially linear
polysaccharide chain to the cell wall. In recent years, a lot
of lipid A analogues have been investigated to ®nd LPS
antagonists as antisepticemia drugs,5±7 and also LPS
agonists as anticancer drugs. Then, a lot of information on
the effect of substitutions in the molecules on the biological
activities has been reported. It has been revealed that the
phosphoric acid group in the 1-position of lipid A can be
exchanged with the carboxymethyl group and dicarboxy-
methyl group without loss of the biological activities of
lipid A.8,9 In addition, the number of fatty acid chains in
lipid A-related compounds appear to be important as to
During our investigation of the biological activities of
compounds related to GLA-60 (2),11 a lipid A-related mono-
saccharide compound which possesses LPS-agonistic
activity, we found that the pyrancarboxylic acid derivative
Keywords: antibacterials; biologically active compounds; glycolipids; lipo-
polysaccharides.
* Corresponding author. Tel.: 181-3-3492-3131; fax: 181-3-5436-8570;
e-mail: shioza@shina.sankyo.co.jp
Figure 1. The structures of E. coli lipid A (1), GLA-60 (2), and the pyran-
carboxylic acid derivative 3.
0040±4020/00/$ - see front matter q 2000 Elsevier Science Ltd. All rights reserved.
PII: S0040-4020(00)00684-0

7692
T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
Figure 2. The structures of lipid A-type pyrancarboxylic acids 4a±c and 5a±c.
3 showed LPS-antagonistic activity.12 Then, expecting that
disaccharide derivatives like lipid A molecules should
have more potent LPS-antagonistic activity, we investigated
the synthesis and biological activities of lipid A-type
(disaccharide) pyrancarboxylic acid derivatives (Fig. 2).13
Herein, we disclose the full details of our study of lipid
A-type pyrancarboxylic acid derivatives.
Results
Synthesis
At ®rst, the starting allyl 2-deoxy-3-O-[(R)-3-tetradecanoyl-
oxytetradecanoyl]-2-(2,2,2-trichloroethoxycarbonylamino)-
a-d-glucopyranoside (6)3b was converted to the three
different 6-substituted glycosyl donors 9, 13, and 15 as
shown in Scheme 1. The 6-hydroxyl group of the diol 6
was selectively protected as a benzyloxycarbonate 7 by
treatment with benzylchloroformate and pyridine according
to the reported method.14 After 4-O-phosphorylation of 7
with diphenyl chlorophosphate and N,N-dimethylamino-
pyridine (DMAP), the allyl group in the anomeric position
of the thus-obtained 8 was deprotected by treatment with
(1,5-cyclooctadiene)bis(methyldiphenylphosphine)iridium(I)
hexa¯uorophosphate in THF and successive hydrolysis with
H2O±I2, affording a 6-O-benzyloxycarbonated intramolecu-
lar hemiacetal 9.15 On the other hand, the synthesis of
6-methoxy and 6-¯uoro derivatives commenced with
selective silylation of the 6-hydroxyl group of 6 with tert-
butyldimethylsilyl chloride and imidazole in DMF to give
the 6-O-monosilylated compound 10. Subsequently, 4-O-
phosphorylation of 10 with diphenyl chlorophosphate and
DMAP, and successive deprotection of the 6-O-silyl group
We ®rst wanted to examine whether the carboxy group
attached directly to the pyran in the anomeric position in
the a-con®guration plays a decisive role in LPS-agonism or
-antagonism of lipid A-type pyrancarboxylic acid deriva-
tives. If not as such, we also wanted to know whether the
phosphate group in the anomeric position of lipid A could be
exchanged to a carboxy group, which would be more stable
than the former, without loss of biological activities.
Further, we tried to investigate the effect of the number of
fatty acid chains in the pyrancarboxylic acid derivatives
upon the biological activities by changing the substituent
in the 20-position in the molecules from (R)-3-(dodecanoyl-
oxy)tetradecanamido groups (4a±c) to acetamido groups
(5a±c). In addition, the effect of a substituent in the
60-position of the compounds was also examined by
comparing the biological activities of 60-hydroxyl (4a,
5a), 60-methoxy (4b, 5b), and 60-¯uoro derivatives (4c, 5c).
Scheme 1. Reagents and conditions: (a) ClCO2Bn, Py, rt, 1 h, 60%; (b) ClPO(OPh)2, DMAP, CH2Cl2, rt, 2 h for 7, 8: 98%, 30 min for 10; (c) [Ir(COD)(P-
MePh2)2]PF6, THF, rt, 1 h, then H2O±I2, 608C, 1 h, 9: 77%, 13: 73%, 15: 79%; (d) t-BuMe2SiCl, imidazole, DMF, rt, 1 h, 84%; (e) 3 M HCl aq., THF, 508C,
1 h, 96% in 2 steps from 10; (f) Me3OBF4, 2,6-di(tert-butyl)-4-methylpyridine, CH2Cl2, rt, 24 h, 68%; (g) DAST, CH2Cl2, 08C, 3 h, 29%.

T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
7693
Scheme 2. Reagents and conditions: (a) PPh3, THF, 2 h, then 28% aq. NH4OH, 608C, 3 h; (b) (R)-3-(benzyloxy)tetradecanoic acid, DCC, CH2Cl2, rt, 15 h,
68% in 2 steps from 16; (c) 4 M HCl, 1,4-dioxane-H2O, 608C, 4 h, then Ph2CN2, THF, 608C, 2 h, 33%; (d) Me2C(OMe)2, p-TsOH, DMF, rt, 3 h, 80%; (e) (R)-
3-(benzyloxy)tetradecanoic acid, DCC, DMAP, CH2Cl2, rt, 16 h, 85%; (f) p-TsOH, MeOH-THF, rt, 3 h, 87%.
with 3 M aqueous hydrochloric acid in THF provided
alcohol 11. After methylation of the 6-hydroxyl group of
11 with trimethyloxonium tetra¯uoroborate and 2,6-di(tert-
butyl)-4-methylpyridine, deprotection of the anomeric allyl
group of the thus-obtained 12 was accomplished by the
same procedure as mentioned above, affording 6-methoxy
hemiacetal 13. Furthermore, after ¯uorination of the
6-hydroxyl group of 11 with diethylaminosulfur tri¯uoride
(DAST), the anomeric allyl group of the thus-obtained 14
was likewise deprotected to yield 6-¯uoro hemiacetal 15.
esteri®cation of the resulting carboxylic acid with diphenyl
diazomethane gave a triol 18. After transformation of 18 to
²
5,7-O-isopropylidene 19 by treatment with 2,2-dimethoxy-
propane and a catalytic amount of p-toluenesulfonic acid
(p-TsOH), esteri®cation of the 4-hydroxyl group of 19
with (R)-3-(benzyloxy)tetradecanoic acid, DCC, and
DMAP yielded compound 20. Then, deprotection of the
isopropylidene group of 20 using p-TsOH as a catalyst in
MeOH-THF furnished glycosyl acceptor 21.
The glycosylation reaction of 9, 13, and 15 with 21 was
conducted by use of Schmidt's imidate method16 (Scheme
3). After conversion of the hemiacetals 9, 13, and 15 to
corresponding trichloroacetimidates with trichloroaceto-
nitrile and a catalytic amount of 1,8-diazabicyclo[5.4.0]-7-
undecene (DBU), successive glycosylation with 21 by use
of trimethylsilyl tri¯uoromethanesulfonate (TMSOTf) as a
catalyst gave rise to only b-oriented pseudodisaccharides
22a, 22b, and 22c, respectively.
Glycosyl acceptor 21 was prepared in accordance with the
reported procedure (Scheme 2).12 The 2-azido group of 2,6-
anhydro-3-azido-3-deoxy-5,7-O-isopropylidene-d-glycero-
d-ido-heptononitrile (16)12 was transformed to the (R)-3-
(benzyloxy)tetradecanamido group of 17 by successive
treatment with triphenylphosphine and aqueous NH4OH,
followed by (R)-3-(benzyloxy)tetradecanoic acid and
dicyclohexyl carbodiimide (DCC). Acidic hydrolysis of
the nitrile 17 with 4 M HCl in dioxane±H2O, and successive
Then, the intermediates 22a, 22b, and 22c were converted
into two kinds of lipid A-type pyrancarboxylic acid deriva-
tives; ones with a (R)-3-(dodecanoyloxy)tetradecanamido
group at the 20-position in the molecules (Scheme 4) similar
to Escherichia coli lipid A, and the others with an acetamido
group at the same position (Scheme 5). After deprotection of
the trichloroethoxycarbonyl (Troc) group at the 20-position
of 22a±c with zinc dust-acetic acid, we treated the thus-
obtained amines with (R)-3-(dodecanoyloxy)tetradecanoic
acid and 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide
³
hydrochloride (EDCI´HCl) to afford 23a±c. Hydrogenoly-
sis of this series of compounds using palladium hydroxide
on carbon as a catalyst completed the deprotection of the
two benzyl ether groups and the diphenylmethyl ester group
of 23a±c, and the benzyloxycarbonyl group of 23a to
furnish 24a±c. Finally, hydrogenolytic deprotection of
each diphenyl phosphate ester group of 24a±c using plati-
num oxide as a catalyst yielded 4a, 4b, and 4c, respectively
²
This hydrolysis step was thoroughly examined, but gave rise to product
18 in 33% yield at most.
Scheme 3. Reagents and conditions: (a) Cl3CCN, DBU, CH2Cl2, 08C,
Ê
30 min; (b) 21, TMSOTf, MS 4A, CH2Cl2, 2508C, 1 h, 22a: 56% in 2
steps from 9, 22b: 57% in 2 steps from 13, 22c: 63% in 2 steps from 15.
³
The given yields of 23a and 23c are the results of only one experiment
each. Higher yields might be obtained with further investigation.

7694
T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
(Scheme 4). We conducted this 2-step reductive deprotec-
tion procedure. So we could avoid the troublesome puri®ca-
tion of the very polar products at the last step. We excluded
the impurities by reduction after the ®rst deprotection step.
On the other hand, after deprotection of the Troc groups of
22a±c by the above-mentioned way, we introduced the
acetamido group at the 20-position with acetic acid and
EDCI´HCl as a condensing agent, affording 25a±c. By the
same procedure as the synthesis of 4a±c, the two benzyl
ether groups, the diphenylmethyl ester group, and the
benzyloxycarbonyl group in the case of 25a were depro-
tected, and then the diphenyl phosphate groups of the
thus-obtained 26a±c were deprotected to furnish 5a, 5b,
and 5c, respectively (Scheme 5).
Biological activity
The biological activities of the newly synthesized
compounds 4a±c and 5a±c were investigated by measuring
TNFa production in human monoblastic U937 cells,
pretreated with TPA, in the absence (LPS agonism) or
presence (LPS antagonism) of LPS (30 ng/ml). As shown
in Figs. 3 and 4, compounds 4a±c, which have six fatty acid
moieties in each molecule, induced the production of TNFa
(LPS-agonistic activity) and showed weak or no inhibitory
activity toward LPS-stimulated TNFa production.
Compound 4c, which showed weaker LPS-agonistic activity
than compounds 4a and 4b, slightly inhibited LPS-induced
TNFa production at a higher concentration of 500 nM. On
the other hand, compounds 5a±c, which have four lipids,
strongly inhibited the production of TNFa induced by LPS
Scheme 5. Reagents and conditions: (a) Zn, AcOH, rt, 5 h; (b) AcOH,
EDCI´HCl, CH2Cl2, rt, 4 h, 25a: 47% in 2 steps from 22a, 25b: 75% in 2
steps from 22b, 25c: 57% in 2 steps from 22c; (c) H2, 20% Pd(OH)2/C,
AcOEt or EtOH, rt, 7±16h, 26a: 63%, 26b: 63%, 26c: 65%; (d) H2, PtO2,
THF, rt, 15±20 h, 5a: 82%, 5b: 99%, 5c: 87%.
in a dose-dependent manner (LPS-antagonistic activity,
Fig. 4), with an IC50 value of 7, 10, 9 nM, respectively,
but did not show any LPS-agonistic activity (Fig. 3).
Discussion
The biological activities of the lipid A-type pyrancarboxylic
acid derivatives synthesized in this work suggest the follow-
ing relationships between the structural features of the
molecules and their biological activities. Firstly, the LPS-
agonistic activity of compounds 4a±c, which have a similar
acyl substitution pattern to E. coli lipid A, and have a
carboxylic acid group directly attached to the anomeric
position of each molecule instead of the phosphate group
in lipid A, revealed that the phosphate group can be replaced
with the carboxy group without loss of biological activity.
The exchangeability of the phosphate group to the stable
carboxy group is useful information for development of
lipid A-type molecules, in which instability of the molecules
in the anomeric position is often encountered.
Secondly, by comparison of the biological activities of
compounds 4a±c with compounds 5a±c, it appears that
the number of lipid chains in the molecule is crucial for
whether the lipid A-type molecule has LPS-agonistic or
-antagonistic activity. Compounds 5a±c, which have four
lipid chains in the molecules, have strong LPS-antagonistic
activities without any LPS-agonistic activity as lipid IVa5
(Fig. 5), which also has four lipid chains in the molecule and
is known as an LPS-antagonist toward human cells. The
importance of the number of lipid chains for biological
Scheme 4. Reagents and conditions: (a) Zn, AcOH, rt, 3 h; (b) (R)-3-(dode-
canoyloxy)tetradecanoic acid, EDCI´HCl, CH2Cl2, rt, 16 h, 23a: 38% in 2
steps from 22a, 23b: 59% in 2 steps from 22b, 23c: 24% in 2 steps from
22c; (c) H2, 20% Pd(OH)2/C, AcOEt or EtOH, rt, 15±20 h, 24a: 57%, 24b:
57%, 24c: 58%; (d) H2, PtO2, THF, rt, 15±20 h, 4a: 88%, 4b: 49%, 4c: 73%.

T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
7695
Figure 3. Production of TNFa from TPA-treated U937 cells stimulated by lipid A-type pyrancarboxylic acid derivatives. In this experiment, the dose-
dependency of the response by LPS (data not shown) and E. coli lipid A17 was con®rmed. As a control, the amounts of TNFa produced by TPA-treated U937
cells stimulated with LPS (30 ng/ml) was 1183 pg/ml.
Figure 4. Inhibition of TNFa release from TPA-treated U937 cells stimulated by LPS (30 ng/ml) in the presence of lipid A-type pyrancarboxylic acid
derivatives.
activities is obvious. This study provides a new acyl substi-
tution pattern for LPS-antagonistic lipid A-related
compounds.
slight inhibition of the TNFa production at a higher
concentrations of LPS-agonistic compound 4c.
Conclusion
Thirdly, there is no signi®cant difference in LPS-antagonis-
tic activities among the 20-acetamido derivatives 5a±c.
Likewise, compounds 4a±c show almost the same level of
LPS-agonistic activity, except for the slight inhibitory
activity of a higher concentration of 4c on LPS-induced
TNFa production, the reason for which was not elucidated.
Therefore, the substituent in the 60-position of the molecules
appears not to be very important for their biological
activities. Further studies are necessary to investigate the
In summary, two kinds of lipid A-type pyrancarboxylic acid
derivatives were synthesized. One of them, having four lipid
chains in the molecules, showed strong LPS-antagonistic
activity. The exchangeability of the phosphate group in
the anomeric-position to a stable carboxy group and the
importance of the lipid chain number suggest an effective
acyl substitution pattern for developing LPS-antagonists.
Experimental
General
1H NMR spectra were recorded using tetramethylsilane as
the internal reference. IR absorption spectra were recorded
on a Jasco IR A-2 spectrophotometer, and mass spectra were
obtained with a JMS-700 mass spectrometer. 3-Nitrobenzyl-
alcohol was used for a matrix on every measurement.
Figure 5. The structure of lipid IVa (27).

7696
T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
Elemental analyses were performed by the Institute of
Science and Technology, Inc. Separation of the compounds
by column chromatography was carried out with silica gel
60 (Merck, 230±400 mesh ASTM).
octadiene)bis(diphenylmethylphosphine)iridium(I) hexa-
¯uorophosphate (33.9 mg, 0.0401 mmol) in THF (5 ml)
was replaced with a hydrogen atmosphere. Stirring for
1 min, the color of the solution turned colorless from red±
brown, then immediately the reaction mixture was replaced
with a nitrogen atmosphere. After stirring for 1 h at room
temperature, H2O (0.5 ml) and I2 (280 mg, 1.10 mmol) were
added to the mixture, and the mixture was stirred for 1 h at
608C. The reaction mixture was quenched with aqueous 5%
Na2SO3, extracted with EtOAc, washed with brine, dried
over Na2SO4, ®ltered, and concentrated in vacuo to give a
mixture. The mixture was puri®ed by silica gel chromato-
graphy. Elution with hexane±EtOAc (4:1) gave 9 (656 mg,
77% yield) as a powder. IR: nmax (KBr) 3344 (broad), 1749,
Allyl 6-O-benzyloxycarbonyl-2-deoxy-3-O-[(R)-3-(tetra-
decanoyloxy)tetradecanoyl]-2-(2,2,2-trichloroethoxycar-
bonylamino)-a-d-glucopyranoside (7). To a solution of
allyl 2-deoxy-3-O-[(R)-3-tetradecanoyloxytetradecanoyl]-
2-(2,2,2-trichloroethoxycarbonylamino)-a-d-glucopyrano-
side (6,3b 438 mg, 0.526 mmol) in THF (30 ml), benzyloxy-
carbonylchloride (125 mg, 0.735 mmol) and pyridine
(62.7 mg, 0.790 mmol) were added. After stirring for 2 h
at room temperature, the reaction mixture was concentrated
in vacuo, diluted with ethyl acetate, washed with 1 M hydro-
chloric acid, saturated aqueous NaHCO3, and brine, dried
over Na2SO4, ®ltered, and concentrated in vacuo to give a
mixture. The mixture was chromatographed on a silica gel
column. Elution with hexane±EtOAc (1:1) gave 7 (304 mg,
60% yield) as a powder. IR: nmax (KBr) 3472, 1748, 1724,
1
1490, 1269 cm21. H NMR (270 MHz, CDCl3) 7.65±7.11
(15H, m), 5.58±5.45 (2H, m), 5.32 (1H, d, Jˆ3.4 Hz), 5.13±
5.01 (3H, m, including 2H, AB-q, Jˆ12.0 Hz, at 5.11,
5.03 ppm), 4.73 (1H, q, Jˆ9.2 Hz), 4.64 (1H, AB-q,
Jˆ12.0 Hz), 4.52 (1H, AB-q, Jˆ12.0 Hz), 4.37±4.18 (3H,
m), 4.03±3.94 (1H, m), 2.45 (2H, d, Jˆ6.3 Hz), 2.16 (2H, t,
Jˆ7.6 Hz), 1.67±1.33 (4H, m), 1.30±1.11 (38H, m), 0.88
(6H, t, Jˆ6.6 Hz). Anal. Calcd for C57H81NO15Cl3P1H2O:
C, 58.24; H, 7.12; N, 1.19; Cl, 9.05; P, 2.63. Found: C,
58.48; H, 7.04; N, 1.23; Cl, 9.32; P, 2.66.
1
1540, 1296 cm21. H NMR (270 MHz, CDCl3) 7.45±7.29
(5H, m), 5.94±5.79 (1H, m), 5.41±5.05 (7H, m, including
2H, s, at 5.18 ppm), 4.90 (1H, d, Jˆ3.5 Hz), 4.76 (1H, AB-
q, Jˆ12.0 Hz), 4.65 (1H, AB-q, Jˆ12.0 Hz), 4.45 (2H, d,
Jˆ3.3 Hz), 4.20±4.11 (1H, m), 4.01±3.85 (3H, m), 3.66±
3.57 (1H, m), 3.40 (1H, d, Jˆ3.9 Hz), 2.56±2.45 (2H, m),
2.29 (2H, t, Jˆ7.5 Hz), 1.68±1.52 (4H, m), 1.30±1.17 (38
H, m), 0.88 (6H, t, Jˆ6.6 Hz). High Resolution MS (FAB,
positive), calcd for C48H76NO12Cl3Na (M1Na)1: 986.4331;
found: 986.4332. Anal. Calcd for C48H76NO12Cl31H2O: C,
58.62; H, 7.99; N, 1.42; Cl, 10.81. Found: C, 58.67; H, 7.90;
N, 1.49; Cl, 11.05.
Allyl 6-O-tert-butyldimethylsilyl-2-deoxy-3-O-[(R)-3-(tetra-
decanoyloxy)tetradecanoyl]-2-(2,2,2-trichloroethoxycar-
bonylamino)-a-d-glucopyranoside (10). To a solution of 6
(2.37 g, 2.86 mmol) in DMF (30 ml), imidazole (398 mg,
5.85 mmol) and tert-butyldimethylsilyl chloride (470 mg,
3.12 mmol) were added. After stirring for 1 h at room
temperature, the reaction mixture was diluted with diethyl
ether, washed with water and brine, dried over Na2SO4,
®ltered, and concentrated in vacuo to give a mixture. The
mixture was chromatographed on a silica gel column.
Elution with hexane±EtOAc (10:1) gave 10 (2.27 g, 84%
yield) as a viscous oil. IR: nmax (neat) 2926, 2856,
Allyl 6-O-benzyloxycarbonyl-2-deoxy-4-O-diphenylphos-
phono-3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-2-
(2,2,2-trichloroethoxycarbonylamino)-a-d-glucopyrano-
side (8). To a solution of 7 (278 mg, 0.288 mmol) in CH2Cl2
(5 ml), DMAP (70.8 mg, 0.58 mmol) and diphenyl chloro-
phosphate (155 mg, 0.577 mmol) were added. After stirring
for 2 h at room temperature, the reaction mixture was
concentrated in vacuo, diluted with ethyl acetate, washed
with saturated aqueous NaHCO3 and brine, and concen-
trated in vacuo to give a mixture. The mixture was puri®ed
using silica gel chromatography. Elution with hexane±
EtOAc (5:1) gave 8 (337 mg, 98% yield) as a viscous oil.
1
1743 cm21. H NMR (400 MHz, CDCl3) 5.94±5.84 (1H,
m), 5.44 (1H, d, Jˆ9.5 Hz, NH), 5.32±5.12 (4H, m), 4.89
(1H, d, Jˆ3.6 Hz), 4.74, 4.68 (2H, AB-q, Jˆ12.5 Hz), 4.19
(1H, dd, Jˆ12.5, 5.1 Hz), 3.99 (1H, dd, Jˆ12.5, 6.6 Hz),
3.94±3.84 (2H, m), 3.73±3.63 (3H, m), 3.38 (1H, broad s,
H), 2.59 (1H, dd, Jˆ15.0, 7.7 Hz), 2.50 (1H, dd, Jˆ15.4,
4.4 Hz), 2.28 (2H, t, Jˆ7.7 Hz), 1.64±1.58 (2H, m), 1.36±
1.20 (40H, m), 0.96±0.86 (15H, m), 0.10 (3H, s), 0.07 (3H,
s). High Resolution MS (FAB, positive), calcd for
IR: nmax (neat) 1751, 1267 cm21 1H NMR (270 MHz,
.
CDCl3) 7.65±7.14 (15H, m), 5.93±5.79 (1H, m), 5.47±
5.38 (2H, m), 5.26 (1H, dd, Jˆ28.2, 1.6 Hz), 5.25 (1H, d,
J ˆ1.1 Hz), 5.13±5.01 (3H, m, including 2H, AB-q,
Jˆ12.1 Hz, at 5.11, 5.01 ppm), 4.79 (1H, AB-q,
Jˆ12.0 Hz), 4.74 (1H, q, Jˆ9.4 Hz), 4.60 (1H, AB-q,
Jˆ12.0 Hz), 4.35±4.11 (3H, m), 4.08±3.95 (3H, m), 2.45
(2H, d, Jˆ6.3 Hz), 2.14 (2H, t, Jˆ7.6 Hz), 1.56±1.40 (4H,
m), 1.33±0.92 (38H, m), 0.88 (6H, t, Jˆ6.6 Hz). High Reso-
lution MS (FAB, positive), calcd for C60H85NO15Cl3PK
(M1K)1: 1234.4359; found: 1234.4381. Anal. Calcd for
C60H85NO15Cl3P: C, 60.17; H, 7.15; N, 1.17; Cl, 8.88; P,
2.59. Found: C, 60.29; H, 6.95; N, 1.30; Cl, 8.68; P, 2.57.
C46H84NO10Cl3SiNa
966.4811.
(M1Na)1:
966.4828;
found:
Allyl 2-deoxy-4-O-diphenylphosphono-3-O-[(R)-3-(tetra-
decanoyloxy)tetradecanoyl]-2-(2,2,2-trichloroethoxycar-
bonylamino)-a-d-glucopyranoside (11). To a solution of
10 (2.27 g, 2.40 mmol) in CH2Cl2 (30 ml), DMAP (589 mg,
4.82 mmol) and diphenyl chlorophosphate (1.30 g,
4.84 mmol) were added. After stirring for 30 min at room
temperature, the reaction mixture was quenched with water,
extracted with EtOAc, washed with brine, dried over
Na2SO4, ®ltered, and concentrated in vacuo to give a
mixture, which was further used without puri®cation. To a
solution of the mixture above obtained in THF (30 ml),
aqueous 3 M hydrochloric acid was added. After stirring
for 1 h at 508C, the reaction mixture was concentrated
in vacuo to give a crude product. The crude product was
6-O-Benzyloxycarbonyl-2-deoxy-4-O-diphenylphos-
phono-3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-2-
(2,2,2-trichloroethoxycarbonylamino)-d-glucopyranose
(9). A solution of 8 (881 mg, 0.735 mmol) and (1,5-cyclo-

T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
7697
puri®ed by silica gel chromatography. Elution with hexane±
EtOAc (2:1) gave 11 (2.46 g, 96% yield) as a viscous oil.
was quenched with aqueous phosphate buffer (pH 7),
extracted with EtOAc, washed with brine, dried over
Na2SO4, ®ltered, and concentrated in vacuo to give a
mixture. The mixture was chromatographed on a silica gel
column. Elution with hexane±EtOAc (2:1) gave 14
(683 mg, 29% yield) as a viscous oil. IR: nmax (KBr)
1
IR: nmax (neat) 3439, 2926, 2855, 1745 cm21. H NMR
(400 MHz, CDCl3) 7.47±7.12 (10H, m), 5.92±5.83 (1H,
m), 5.47 (1H, t, Jˆ9.6 Hz), 5.39 (1H, d, Jˆ9.7 Hz), 5.31
(1H, dd, Jˆ17.3, 1.3 Hz), 5.27±5.23 (1H, m), 5.13 (1H, t,
Jˆ6.3 Hz), 4.97 (1H, d, Jˆ3.5 Hz), 4.80±4.73(1H, m), 4.76
(1H, AB-q, Jˆ12.0 Hz), 4.65 (1H, AB-q, Jˆ12.0 Hz),
4.21±4.16 (1H, m), 4.08±3.98 (2H, m), 3.75 (1H, d,
Jˆ9.8 Hz), 3.64±3.62 (2H, m), 3.32±3.28 (1H, m), 2.42±
2.40 (2H, m), 2.18 (2H, t, Jˆ7.6 Hz), 1.60±1.40 (2H, m),
1.38±1.16 (40H, m), 0.88 (3H, t, Jˆ6.8 Hz). High Resolu-
tion MS (FAB, positive), calcd for C52H79NO13Cl3PNa
(M1Na)1: 1084.4252; found: 1084.4258.
1747, 1591, 1516, 1491 cm21
.
1H NMR (400 MHz,
CDCl3) 7.36±7.16 (10H, m), 5.91±5.84 (1H, m), 5.48±
5.40 (2H, m), 5.34±5.24 (2H, m), 5.13±5.10 (1H, m), 4.98
(1H, d, Jˆ3.7 Hz), 4.80 (1H, AB-q, Jˆ12.5 Hz), 4.72 (1H,
q, Jˆ9.5 Hz), 4.58 (1H, AB-q, Jˆ12.5 Hz), 4.46 (2H, dd,
Jˆ47.2, 2.6 Hz), 4.23±4.18 (1H, m), 4.06±3.95 (3H, m),
2.45 (2H, d, Jˆ6.6 Hz), 2.14 (2H, t, Jˆ7.3 Hz), 1.60±1.17
(44H, m), 0.88 (6H, t, Jˆ7.0 Hz). High Resolution MS
(FAB, positive), calcd for C52H78NO12Cl3FPNa (M1Na)1:
1086.4209; found: 1086.4207. Anal. Calcd for
C52H78NO12Cl3FP: C, 58.62; H, 7.38; N, 1.32; F, 1.78; Cl,
9.98; P, 2.91. Found: C, 58.65; H, 7.41; N, 1.24; F, 1.67; Cl,
9.89; P, 3.04.
Allyl 2-deoxy-4-O-diphenylphosphono-6-O-methyl-3-O-
[(R)-3-(tetradecanoyloxy)tetradecanoyl]-2-(2,2,2-tri-
chloroethoxycarbonylamino)-a-d-glucopyranoside (12).
To a solution of 11 (13.1 mg, 0.012 mmol) in CH2Cl2
(1 ml), a solution of trimethyloxonium tetra¯uoroborate
(36.0 mg, 0.243 mmol) and 2,6-di(tert-butyl)-4-methyl-
pyridine (9.1 mg, 0.044 mmol) were added. After stirring
for 24 h at room temperature, the reaction mixture was
quenched with water, extracted with EtOAc, washed with
brine, dried over Na2SO4, ®ltered, and concentrated in
vacuo to give a mixture. The mixture was chromatographed
on a silica gel column. Elution with hexane±EtOAc (5:1)
gave 12 (9.0 mg, 68% yield) as a viscous oil. IR: nmax (KBr)
2,6-Deoxy-4-O-diphenylphosphono-6-¯uoro-3-O-[(R)-3-
(tetradecanoyloxy)tetradecanoyl]-2-(2,2,2-trichloro-
ethoxycarbonylamino)-d-glucopyranose (15). Compound
14 (683.4 mg, 0.641 mmol) was treated as described in the
formation of 9 from 8 to give 15 (518 mg, 79% yield) as a
powder. IR: nmax (KBr) 3424 (broad), 1728, 1524 cm21. 1H
NMR (270 MHz, CDCl3) 7.36±7.17 (10H, m), 5.66 (1H, d,
Jˆ9.5 Hz), 5.55 (1H, t, Jˆ9.9 Hz), 5.37 (1H, d, Jˆ3.7 Hz),
5.13±5.09 (1H, m), 4.75 (1H, AB-q, Jˆ11.7 Hz), 4.72 (1H,
q, Jˆ9.5 Hz), 4.64 (1H, AB-q, Jˆ11.7 Hz), 4.46 (2H, d,
Jˆ47.6 Hz), 4.26 (1H, dd, Jˆ12.5, 6.2 Hz), 4.03±3.97
(1H, m), 2.45 (2H, d, Jˆ6.6 Hz), 2.15 (2H, t, Jˆ7.3 Hz),
1.53±1.20 (42 H, m), 0.88 (6H, t, Jˆ7.0 Hz). High Resolu-
tion MS (FAB, positive), calcd for C49H75NO12Cl3FP
(M1H)1: 1024.4076; found: 1024.4027. Anal. Calcd for
C49H74NO12Cl3FP: C, 57.39; H, 7.27; N, 1.37; F, 1.85; Cl,
10.37; P, 3.02. Found: C, 57.11; H, 7.05; N, 1.42; F, 1.96;
Cl, 10.74; P, 2.90.
1
1748, 1516 cm21. H NMR (270 MHz, CDCl3) 7.37±7.17
(10H, m), 5.96±5.81 (1H, m), 5.48±5.22 (4H, m), 5.16±
5.00 (1H, m), 4.96 (1H, d, Jˆ3.5 Hz), 4.84±4.73 (2H, m,
including 1H, AB-q, Jˆ11.8 Hz at 4.78 ppm), 4.61 (1H,
AB-q, Jˆ11.8 Hz), 4.25±4.18 (1H, m), 4.05±3.90 (3H,
m), 3.52±3.42 (2H, m), 3.20 (3H, s), 2.45 (2H, d,
Jˆ6.3 Hz), 2.13 (2H, t, Jˆ7.5 Hz), 1.61±1.13 (42 H, m),
0.88 (6H, t, Jˆ6.6 Hz). High Resolution MS (FAB, posi-
tive), calcd for C53H81NO13Cl3PNa (M1Na)1: 1098.4409;
found: 1098.4390. Anal. Calcd for C53H81NO13Cl3P: C,
59.08; H, 7.58; N, 1.30; Cl, 9.87; P, 2.87. Found: C,
58.40; H, 7.71; N, 1.26; Cl, 10.15; P, 2.84.
2,6-Anhydro-3-[(R)-3-(benzyloxy)tetradecanamido]-3-
deoxy-5,7-O-isopropylidene-d-glycero-d-ido-heptono-
nitrile (17). To a solution of 2,6-anhydro-3-azido-3-deoxy-
5,7-O-isopropylidene-d-glycero-d-ido-heptononitrile (16,12
10.1 g, 39.6 mmol) in THF (200 ml), triphenylphosphine
(11.4 g, 43.6 mmol) was added at room temperature. After
stirring for 2 h, 28% aqueous NH4OH was added to the
reaction mixture, which was further stirred at 608C for
3 h. Then, the reaction mixture was concentrated in vacuo,
and coevaporated with toluene to remove H2O. The
obtained precipitate was dissolved in CH2Cl2 (100 ml),
and to the solution, a solution of (R)-3-(benzyloxy)tetra-
decanoic acid (13.3 g, 39.8 mmol) in CH2Cl2 (70 ml) and
a solution of DCC (8.25 g, 40.0 mmol) in CH2Cl2 (30 ml)
were added. After stirring for 15 h, the resulting white pre-
cipitate was removed by ®ltration, and the solution was
concentrated in vacuo to give a mixture. The mixture was
puri®ed using silica gel chromatography. Elution with
hexane±EtOAc (2:1) gave 17 (14.6 g, 68% yield) as an
2-Deoxy-4-O-diphenylphosphono-6-O-methyl-3-O-[(R)-
3-(tetradecanoyloxy)tetradecanoyl]-2-(2,2,2-trichloro-
ethoxycarbonylamino)-d-glucopyranose (13). Compound
12 (898 mg, 0.833 mmol) was treated as described in the
formation of 9 from 8 to give 13 (629 mg, 73% yield) as a
powder. IR: nmax (KBr) 3399 (broad), 1728, 1525 cm21. 1H
NMR (270 MHz, CDCl3) 7.37±7.18 (10H, m), 5.66 (1H, d,
Jˆ9.5 Hz), 5.50 (1H, dd, Jˆ10.3, 9.8 Hz), 5.32 (1H, d,
Jˆ3.1 Hz), 5.15±5.06 (1H, m), 4.79±4.62 (3H, m), 4.32
(1H, broad s, OH), 4.22±4.18 (1H, m), 3.98 (1H, dt,
Jdˆ3.1 Hz, Jtˆ10.1 Hz), 3.52±3.37 (2H, m), 3.20 (3H, s),
2.44 (2H, d, Jˆ6.1 Hz), 2.15 (2H, t, Jˆ7.6 Hz), 1.55±1.11
(42 H, m), 0.88 (6H, t, Jˆ6.6 Hz). High Resolution MS
(FAB, positive), calcd for C50H78NO13Cl3P (M1H)1:
1036.4295; found: 1036.4297.
Allyl 2,6-deoxy-4-O-diphenylphosphono-6-¯uoro-3-O-[(R)-
3-(tetradecanoyloxy)tetradecanoyl]-2-(2,2,2-trichloro-
ethoxycarbonylamino)-a-d-glucopyranoside (14). To a
solution of 11 (2.38 g, 2.24 mmol) in CH2Cl2 (100 ml),
diethylaminosulfurtri¯uoride (2 ml, 15.1 mmol) was added
at 2358C. After stirring for 3 h at 08C, the reaction mixture
amorphous ®lm. IR: nmax (neat) 3418, 1644, 1628 cm21
.
1H NMR (270 MHz, CDCl3) 7.40±7.32 (10H, m), 7.03
(1H, d, Jˆ6.0 Hz), 5.20 (1H, d, Jˆ6.0 Hz), 4.64 (1H, AB-
q, Jˆ11.2 Hz), 4.54 (1H, AB-q, Jˆ11.2 Hz), 4.06 (1H, m),
3.91 (1H, dd, Jˆ9.7, 4.3 Hz), 3.82 (1H, m), 3.72 (1H, t,

7698
T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
Jˆ10.4±9.8 Hz), 3.65 (1H, m), 3.55±3.44 (2H, m), 2.58±
2.41 (2H, m), 1.74±1.50 (2H, m), 1.49 (3H, s), 1.43 (3H, s),
1.26 (18H, m), 0.88 (3H, t, Jˆ7.0±6.2 Hz). High Resolution
MS (FAB, positive), calcd for C31H49N2O6 (M1H)1:
545.3591; found: 545.3579.
hexane±EtOAc (10:1) gave 20 (747 mg, 85% yield) as a
powder. IR: nmax (KBr) 2920, 1740 cm21 1H NMR
.
(270 MHz, CDCl3) 7.54±7.08 (20H, m), 6.85 (1H, s), 6.56
(1H, d, Jˆ8.8 Hz), 5.40 (1H, t, Jˆ9.8 Hz), 4.60±4.37 (6H,
m), 3.81±3.63 (5H, m), 3.31±3.22 (1H, m), 2.65 (1H, dd,
Jˆ15.1, 6.4 Hz), 2.39 (1H, dd, Jˆ15.3, 5.9 Hz), 2.20±2.04
(2H, m), 1.66±1.15 (46H, m), 0.88 (6H, t, Jˆ6.8±6.4 Hz).
High Resolution MS (FAB, positive), calcd for
C65H91NO10Na (M1Na)1: 1068.6541; found: 1068.6543.
Anal. Calcd for C65H91NO10: C, 74.61; H, 8.77; N, 1.34.
Found: C, 74.89; H, 8.68; N, 1.28.
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-3-deoxy-d-glycero-d-ido-heptonate (18). To
a solution of nitrile 17 (3.44 g, 3.99 mmol) in dioxane
(22 ml)-H2O (2.2 ml), 4 M hydrogen chloride dioxane solu-
tion was added. After stirring for 4 h at 608C, the reaction
mixture was concentrated in vacuo, giving a precipitate. The
precipitate was dissolved in dimethylformamide (DMF,
16 ml), treated with diphenyldiazomethane (3.10 g,
15.9 mmol), and stirred for 2 h at 608C. The reaction
mixture was concentrated in vacuo to give a mixture. The
mixture was puri®ed using silica gel chromatography.
Elution with EtOAc, and then EtOAc-MeOH (10:1) gave
18 (897 mg, 33% yield) as a powder. IR: nmax (KBr) 3324,
1734, 1645, 1538 cm21. 1H NMR (270 MHz, CDCl3) 7.41±
7.21 (15H, m), 6.98 (1H, d, Jˆ9.1 Hz), 6.82 (1H, s), 4.55
(1H, d, Jˆ5.7 Hz), 4.40 (2H, AB-q, Jˆ11.5 Hz), 4.33 (1H,
m), 3.81±3.57 (6H, m), 3.28 (1H, m), 2.87 (1H, broad s),
2.42±2.27 (2H, m), 1.65±1.40 (2H, m), 1.30±1.25 (18H,
m), 0.88 (3H, t, Jˆ6.6 Hz). High Resolution MS (FAB,
positive), calcd for C41H56NO8 (M1H)1: 690.4006; found:
690.4016. Anal. Calcd for C41H55NO8: C, 71.37; H, 8.04; N,
2.03. Found: C, 71.25; H, 8.17; N, 2.00.
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-4-O-[(R)-3-(benzyloxy)tetradecanoyl]-3-
deoxy-d-glycero-d-ido-heptonate (21). To a solution of
isopropylidene 20 (1.09 g, 1.04 mmol) in THF (15 ml) and
MeOH (15 ml), p-TsOH (235 mg) was added at 248C. After
stirring for 3 h at 248C, the reaction mixture was concen-
trated in vacuo, and puri®ed with silica gel chromatography.
Elution with hexane±EtOAc (2:1) gave 21 (906 mg, 87%
yield) as a powder. IR: nmax (KBr) 3350, 1733, 1649,
1
1536 cm21. H NMR (270 MHz, CDCl3) 7.33±7.08 (20H,
m), 6.85 (1H, s), 6.61 (1H, d, Jˆ8.8 Hz), 5.27 (1H, t,
Jˆ9.9 Hz), 4.63 (1H, d, Jˆ5.8 Hz), 4.50±4.32 (5H, m,
including two 2H, s, at 4.48 and 4.37 ppm), 3.99±3.80
(1H, m), 3.73±3.58 (5H, m), 3.37±3.30 (1H, m), 3.14±
2.76 (1H, broad s), 2.62±2.42 (2H, m), 2.22±2.14 (2H,
m), 1.69±1.14 (40H, m), 0.88 (6H, t, Jˆ6.4 Hz). High Reso-
lution MS (FAB, positive), calcd for C62H88NO10 (M1H)1:
1006.6408; found: 1006.6397. Anal. Calcd for C62H87NO10:
C, 74.00; H, 8.71; N, 1.39. Found: C, 74.19; H, 8.67; N,
1.40.
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-3-deoxy-5,7-O-isopropylidene-d-glycero-d-
ido-heptonate (19). A solution of triol 18 (781 mg,
1.13 mmol) and p-TsOH (52 mg) in DMF (8 ml) and 2,2-
dimethoxypropane (8 ml) was stirred for 3 h at room
temperature, diluted with diethyl ether, extracted with
10% aqueous NaHCO3, washed with brine, and dried over
MgSO4. The mixture was ®ltered, concentrated in vacuo,
and puri®ed with silica gel chromatography. Elution with
hexane±EtOAc (2:1) gave 19 (660 mg, 80% yield) as a
Diphenylmethyl 2,6-anhydro-7-O-[6-O-benzyloxycar-
bonyl-2-deoxy-4-O-diphenylphosphono-3-O-[(R)-3-(tetra-
decanoyloxy)tetradecanoyl]-2-(2,2,2-trichloroethoxy-
carbonylamino)-b-d-glucopyranosyl]-3-[(R)-3-(benzyl-
oxy)tetradecanamido]-4-O-[(R)-3-(benzyloxy)tetra-
decanoyl]-3-deoxy-d-glycero-d-ido-heptonate (22a). To a
solution of 9 (249 mg, 0.215 mmol) and trichloroaceto-
nitrile (200 mg, 2.00 mmol) in CH2Cl2 (3.0 ml), a solution
of DBU (6.8 mg, 0.0446 mmol) in CH2Cl2 (1.5 ml) was
added at 08C under nitrogen. After stirring for 30 min at
08C, the reaction mixture was concentrated in vacuo, and
rapidly chromatographed with hexane±EtOAc (2:1). The
eluent was concentrated in vacuo to give a crude imidate,
which was immediately used for the next reaction without
further puri®cation. To a suspension in CH2Cl2 (4.0 ml) of
the thus-obtained imidate, diol 21 (217 mg, 0.216 mmol)
1
powder. IR: nmax (KBr) 3376, 1728, 1640 cm21. H NMR
(270 MHz, CDCl3) 7.38±7.24 (15H, m), 6.84 (1H, s), 6.83
(1H, d, Jˆ6.9 Hz), 4.56 (1H, d, Jˆ5.9 Hz), 4.51±4.35 (3H,
m, including 1H, d, Jˆ5.9 Hz, at 4.41 ppm), 3.86±3.62 (4H,
m), 3.56 (1H, t, Jˆ9.3 Hz), 3.26±3.17 (1H, m), 2.54 (1H, d,
Jˆ2.8 Hz), 2.44±2.29 (2H, m), 1.65±1.36 (8H, m, contain-
ing two 3H, s, at 1.48 and 1.37 ppm), 1.29±1.23 (18H, m),
0.88 (3H, t, Jˆ6.8±6.4 Hz). High Resolution MS (FAB,
positive), calcd for C44H60NO8 (M1H)1: 730.4319; found:
730.4307. Anal. Calcd for C44H59NO8: C, 72.40; H, 8.15; N,
1.92. Found: C, 71.95; H, 7.85; N, 2.22.
Ê
and molecular sieves 4A dried in advance, TMSOTf
(4.8 mg, 0.0217 mmol) in CH2Cl2 (50 ml) was added at
2508C. After stirring for 1 h at 2508C, the mixture was
quenched with saturated NaHCO3 aqueous solution,
extracted with EtOAc, and the extract was washed with
brine, dried over Na2SO4, ®ltered, and concentrated in
vacuo to give a mixture. The mixture was puri®ed with
silica gel chromatography. Elution with hexane±EtOAc
(3:1) gave 22a (289 mg, 56% yield) as a powder. IR: nmax
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-4-O-[(R)-3-(benzyloxy)tetradecanoyl]-3-
deoxy-5,7-O-isopropylidene-d-glycero-d-ido-heptonate
(20). To a solution of alcohol 19 (610 mg, 0.835 mmol) in
CH2Cl2 (8 ml), (R)-3-(benzyloxy)tetradecanoic acid
(317 mg, 0.947 mmol), DCC (207 mg, 1.00 mmol), and
DMAP (124 mg, 1.01 mmol) were added at 248C. After
stirring for 16 h at 248C, the reaction mixture was diluted
with ethyl acetate, washed with 0.1 M aqueous HCl, satu-
rated aqueous NaHCO3, and brine in order, and dried over
MgSO4. The mixture was ®ltered, concentrated in vacuo,
and puri®ed using silica gel chromatography. Elution with
1
(KBr) 3343, 1735, 1531, 1268 cm21. H NMR (270 MHz,
CDCl3) 7.38±7.11 (35H, m), 6.84 (1H, s), 6.51 (1H, d,
Jˆ8.9 Hz), 5.71 (1H, d, Jˆ6.8 Hz), 5.55±5.48 (1H, m),
5.32±5.01 (4H, m, including 1H, AB-q, Jˆ12.3 Hz at
5.03 ppm and 1H, AB-q, Jˆ12.3 Hz at 5.12 ppm), 4.99

T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
7699
(1H, d, Jˆ9.7 Hz), 4.75±4.61 (4H, m), 4.57±4.29 (6H, m,
including 1H, s at 4.33 ppm, 1H, s, at 4.35 ppm, 1H, s at
4.48 ppm, and 1H, s at 4.49 ppm), 4.22±4.16 (1H, m),
4.01±3.95 (1H, m), 3.85±3.80 (1H, m), 3.74±3.54 (4H,
m), 3.45±3.34 (2H, m), 2.82 (1H, broad s), 2.64±2.13
(8H, m), 1.67±1.11 (82H, m), 0.88 (12H, t, Jˆ6.5 Hz).
High Resolution MS (FAB, positive), calcd for
C119H166N2O24Cl3PNa (M1Na)1: 2166.0531; found:
2166.0549. Anal. Calcd for C119H166N2O24Cl3P: C, 66.61;
H, 7.80; N, 1.31; Cl, 4.96; P, 1.44. Found: C, 66.14; H, 7.59;
N, 1.29; Cl, 5.16; P, 1.71.
22a (174 mg, 0.080 mmol) in acetic acid (5.0 ml), zinc dust
(745 mg, 11.4 mmol) was added. After vigorously stirring
for 3 h at room temperature, the solution was ®ltered
through Celite to remove the zinc dust. The ®ltrate was
concentrated in vacuo to give a residue, which was
dissolved in CHCl3, washed with saturated aqueous
NaHCO3 solution, dried with MgSO4, and ®ltered. The
®ltrate was concentrated in vacuo, and dissolved in
CH2Cl2 (2.0 ml). To this solution, were added (R)-3-(dode-
canoyloxy)tetradecanoic acid (34.7 mg, 0.081 mmol) and
EDCI´HCl (31.0 mg, 0.162 mmol) at room temperature,
and the mixture was allowed to stand for 16 h at room
temperature. Then, the reaction mixture was concentrated
in vacuo, and chromatographed on a silica gel column
eluted with hexane±EtOAc (3:1) to give 23a (71.8 mg,
38%) as a powder. IR: nmax (KBr) 3341, 1732, 1651,
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-4-O-[(R)-3-(benzyloxy)tetradecanoyl]-3-
deoxy-7-O-[2-deoxy-4-O-diphenylphosphono-6-O-methyl-
3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-2-(2,2,2-
trichloroethoxycarbonylamino)-b-d-glucopyranosyl]-d-
glycero-d-ido-heptonate (22b). Compound 13 (21.5 mg,
0.021 mmol) was treated as described in the formation of
22a from 9 to give 22b (23.3 mg, 57% yield) as a powder.
1
1493 cm21. H NMR (270 MHz, CDCl3) 7.58±7.10 (35H,
m), 6.83 (1H, s), 6.50 (1H, d, Jˆ8.8 Hz), 6.42 (1H, d,
Jˆ7.0 Hz), 5.51±5.47 (1H, m), 5.32±5.28 (1H, m), 5.18±
4.99 (5H, m, containing 1H, AB-q, Jˆ12.2 Hz at 5.11 ppm,
1H, AB-q, Jˆ12.2 Hz at 5.01 ppm), 4.64±4.29 (9H, m),
4.24±4.17 (1H, m), 4.10±3.97 (1H, m), 3.83±3.60 (5H,
m), 3.43±3.39 (1H, m), 2.61±2.08 (12H, m), 1.66±1.14
(120H, m), 0.88 (18H, t, Jˆ6.2 Hz). High Resolution MS
(FAB, positive), calcd for C142H213N2O25PNa (M1Na)1:
2400.5093; found: 2400.5078. Anal. Calcd for
C142H213N2O25P1H2O: C, 71.15; H, 9.04; N, 1.17; P, 1.29.
Found: C, 71.10; H, 8.78; N, 1.13; P, 1.04.
1
IR: nmax (KBr) 3550±3360, 1738, 1531 cm21. H NMR
(270 MHz, CDCl3) 7.36±7.15 (30H, m), 6.84 (1H, s), 6.50
(1H, d, Jˆ8.8 Hz), 5.62 (1H, d, Jˆ6.6 Hz), 5.47 (1H, t,
Jˆ9.8 Hz), 5.31±5.17 (2H, m), 4.92 (1H, d, Jˆ8.3 Hz),
4.75±4.61 (4H, m), 4.50±4.34 (5H, m), 4.01±3.97 (1H,
m), 3.86±3.73 (2H, m), 3.65±3.47 (3H, m), 3.44±3.38
(4H, m), 3.20 (3H, s), 2.65±2.14 (8H, m), 1.66±1.11
(94H, m), 0.88 (12H, t, Jˆ6.1±6.9 Hz). High Resolution
MS (FAB, positive), calcd for C112H162N2O22Cl3PNa
(M1Na)1: 2046.0320; found: 2046.0288. Anal. Calcd for
C112H162N2O22Cl3P: C, 66.40; H, 8.06; N, 1.38; Cl, 5.25; P,
1.53. Found: C, 66.32; H, 7.85; N, 1.37; Cl, 5.41; P, 1.76.
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-4-O-[(R)-3-(benzyloxy)tetradecanoyl]-3-
deoxy-7-O-[2-deoxy-4-O-diphenylphosphono-2-[(R)-3-
(dodecanoyloxy)tetradecanamido]-6-O-methyl-3-O-[(R)-
3-(tetradecanoyloxy)tetradecanoyl]-b-d-glucopyranosyl]-
d-glycero-d-ido-heptonate (23b). Compound 22b (189 mg,
0.093 mmol) was treated as described in the formation of
23a from 22a to give 23b (123 mg, 59%) as a powder. IR:
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-4-O-[(R)-3-(benzyloxy)tetradecanoyl]-3-
deoxy-7-O-[2,6-deoxy-4-O-diphenylphosphono-6-¯uoro-
3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-2-(2,2,2-
trichloroethoxycarbonylamino)-b-d-glucopyranosyl]-d-
glycero-d-ido-heptonate (22c). Compound 15 (312 mg,
0.304 mmol) was treated as described in the formation of
22a from 9 to give 22c (385 mg, 63% yield) as a powder. IR:
1
nmax (KBr) 1736, 1659 cm21. H NMR (270 MHz, CDCl3)
7.42±7.14 (30H, m), 6.83 (1H, s), 6.50 (1H, d, Jˆ9.3 Hz),
6.35 (1H, d, Jˆ7.3 Hz), 5.43 (1H, t, Jˆ9.3 Hz), 5.31 (1H,
dd, Jˆ10.5, 9.2 Hz), 5.17±5.12 (2H, m), 5.04 (1H, d,
Jˆ8.6 Hz), 4.68±4.34 (6H, m), 3.98±3.40 (11H, m),
3.22±3.18 (1H, broad s), 3.18 (3H, s), 2.68±2.04 (12H,
m), 1.74±1.23 (120H, m), 0.88 (18H, t, Jˆ6.3 Hz). High
1
nmax (KBr) 3424, 1732, 1654, 1591, 1491, 1189 cm21. H
NMR (400 MHz, CDCl3) 7.35±7.14 (30H, m), 6.84 (1H, s),
6.52 (1H, d, Jˆ8.8 Hz), 5.75±5.69 (1H, m), 5.58±5.50 (1H,
m), 5.29±5.16 (2H, m), 5.02 (1H, d, Jˆ8.1 Hz), 4.77±4.70
(2H, m), 4.66±4.57 (3H, m), 4.52±4.38 (5H, m), 4.34 (1H,
d, Jˆ5.1 Hz), 4.01 (1H, d, Jˆ7.4 Hz), 3.84±3.81 (1H, m),
3.73±3.59 (4H, m), 3.48±3.43 (1H, m), 3.39±3.32 (1H, m),
2.77 (1H, broad s), 2.60 (1H, dd, Jˆ8.1, 15.4 Hz), 2.45 (1H,
d, Jˆ5.1 Hz), 2.41 (1H, d, Jˆ4.4 Hz), 2.32 (1H, dd, Jˆ8.1,
15.4 Hz), 2.24±2.13 (4H, m), 1.30±1.08 (82H, m), 0.88
(12H, t, Jˆ6.6 Hz). High Resolution MS (FAB, positive),
calcd for C111H159N2O21Cl3FPNa (M1Na)1: 2034.0120;
found: 2034.0135. Anal. Calcd for C111H159N2O21Cl3FP:
C, 66.20; H, 7.96; N, 1.39; Cl, 5.28; F, 0.94; P, 1.54.
Found: C, 65.81; H, 7.69; N, 1.49; Cl, 5.32; F, 0.97; P, 1.29.
Resolution
C135H209N2O23PNa
MS
(FAB,
positive),
calcd
for
found:
(M1Na)1:
2280.4882;
2280.4871. Anal. Calcd for C135H209N2O23P1H2O: C,
71.21; H, 9.34; N, 1.23; P, 1.36. Found: C, 71.21; H, 9.10;
N, 1.28; P, 1.13.
Diphenylmethyl 2,6-anhydro-3-[(R)-3-(benzyloxy)tetra-
decanamido]-4-O-[(R)-3-(benzyloxy)tetradecanoyl]-3-
deoxy-7-O-[2,6-deoxy-4-O-diphenylphosphono-2-[(R)-3-
(dodecanoyloxy)tetradecanamido]-6-¯uoro-3-O-[(R)-3-
(tetradecanoyloxy)tetradecanoyl]-b-d-glucopyranosyl]-
d-glycero-d-ido-heptonate (23c). Compound 22c (123 mg,
0.061 mmol) was treated as described in the formation of
23a from 22a to give 23c (33.1 mg, 24%) as a powder. IR:
Diphenylmethyl 2,6-anhydro-7-O-[6-O-benzyloxycarbonyl-
2-deoxy-4-O-diphenylphosphono-2-[(R)-3-(dodecanoyloxy)-
tetradecanamido]-3-O-[(R)-3-(tetradecanoyloxy)tetra-
decanoyl]-b-d-glucopyranosyl]-3-[(R)-3-(benzyloxy)-
tetradecanamido]-4-O-[(R)-3-(benzyloxy)tetradecanoyl]-
3-deoxy-d-glycero-d-ido-heptonate (23a). To a solution of
1
nmax (KBr) 1737, 1666, 1193 cm21. H NMR (400 MHz,
CDCl3) 7.33±7.13 (30H, m), 6.84 (1H, s), 6.51 (1H, d,
Jˆ8.8 Hz), 6.44 (1H, d, Jˆ6.6 Hz), 5.54 (1H, t, Jˆ
9.5 Hz), 5.31 (1H, t, Jˆ9.5 Hz), 5.20 (1H, d, Jˆ8.1 Hz),
5.19±5.13 (2H, m), 4.64 (1H, d, Jˆ5.1 Hz), 4.62±4.31

7700
T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
(7H, m), 4.02 (1H, d, Jˆ9.5 Hz), 3.89±3.81 (1H, m), 3.78±
3.60 (5H, m), 3.48±3.44 (2H, m), 2.62 (1H, q, Jˆ7.7 Hz),
2.47±2.10 (11H, m), 1.81±1.25 (120H, m), 0.88 (18H, t,
Jˆ7.3±5.1 Hz). High Resolution MS (FAB, positive),
calcd for C134H206N2O22FPNa (M1Na)1: 2268.4682.
Found: 2268.4746.
4-O-[(R)-3-(hydroxy)tetradecanoyl]-d-glycero-d-ido-
heptonic acid (24c). To a solution of 23c (16.5 mg,
0.007 mmol) in ethanol (2.0 ml), 20% Pd(OH)2 on carbon
(19.0 mg) was added, stirred for 20 h under hydrogen at
atmospheric pressure at room temperature, and then, ®ltered
through Celite. The ®ltrate was concentrated in vacuo, and
separated by preparative thin layer chromatography
(CHCl3±MeOH, 8:1). The part containing a product was
eluted with CHCl3±MeOH (5:1) to give a mixture of 24c
contaminated with silica gel. The mixture was dissolved in
CHCl3±MeOH±0.1 M HCl aq. (5:10:4) to remove the silica
gel, and extracted with CHCl3. The chloroform layer was
concentrated in vacuo to give 24c (8.1 mg, 58%) as an
2,6-Anhydro-3-deoxy-7-O-[2-deoxy-4-O-diphenylphos-
phono-2-[(R)-3-(dodecanoyloxy)tetradecanamido]-3-O-
[(R)-3-(tetradecanoyloxy)tetradecanoyl]-b-d-glucopyra-
nosyl]-3-[(R)-3-(hydroxy)tetradecanamido]-4-O-[(R)-3-
(hydroxy)tetradecanoyl]-d-glycero-d-ido-heptonic acid
(24a). To a solution of 23a (73.4 mg, 0.031 mmol) in
ethyl acetate (3.0 ml), 20% Pd(OH)2 on carbon (73.6 mg)
was added, stirred for 15 h under hydrogen at atmospheric
pressure at room temperature, and then, ®ltered through
Celite. The ®ltrate was concentrated in vacuo, and separated
by preparative thin layer chromatography (CHCl3±MeOH,
5:1). The part containing a product was eluted with CHCl3±
MeOH (5:1) to give a mixture of 24a contaminated with
silica gel. The mixture was dissolved in CHCl3±MeOH±
0.1 M HCl aq. (5:10:4) to remove the silica gel, and
extracted with CHCl3. The chloroform layer was concen-
trated in vacuo to give 24a (33.5 mg, 57%) as an amorphous
®lm. IR: nmax (KBr) 3372, 1736, 1664, 1490, 1189 cm21. 1H
NMR (400 MHz, CD3OD±CDCl3(10:1)) 7.41±7.32 (4H,
m), 7.26±7.17 (6H, m), 5.44 (1H, dd, Jˆ10.3, 9.5 Hz),
5.24 (1H, dd, Jˆ10.3, 9.5 Hz), 5.27±5.18 (2H, m), 4.91
(1H, d, Jˆ8.1 Hz), 4.73 (1H, q, Jˆ8.8 Hz), 4.25 (1H, dd,
Jˆ5.9, 5.1 Hz), 4.15 (1H, d, Jˆ5.9 Hz), 4.06 (1H, d,
Jˆ11.0 Hz), 3.68±3.58 (2H, m), 3.45 (1H, t, Jˆ9.5 Hz),
2.59±2.40 (6H, m), 2.38±2.22 (4H, m), 2.14 (2H, t,
Jˆ7.3 Hz), 1.68±1.20 (120H, m), 0.90 (18H, t, Jˆ7.3±
6.6 Hz). High Resolution MS (FAB, positive), calcd for
amorphous ®lm. IR: nmax (KBr) 1736, 1661, 1189 cm21
.
1H NMR (400 MHz, CD3OD±CDCl3(10:1)) 7.42±7.35
(4H, m), 7.26±7.16 (6H, m), 5.48 (1H, t, Jˆ9.9 Hz),
5.29±5.17 (3H, m), 5.03 (1H, d, Jˆ8.1 Hz), 4.72 (1H, t,
Jˆ9.2 Hz), 4.59±4.47 (2H, m), 4.42±4.33 (1H, m), 4.33±
4.23 (1H, m), 4.22±4.12 (1H, m), 4.07 (1H, d, Jˆ11.7 Hz),
4.03±3.96 (1H, m), 3.95±3.78 (3H, m), 3.77±3.70 (1H, m),
3.43 (1H, t, Jˆ9.9 Hz), 2.61±2.42 (6H, m), 2.36±2.25 (4H,
m), 2.18±2.12 (2H, m), 1.68±1.20 (120H, m), 0.90 (18H, t,
Jˆ7.3±5.9 Hz). High Resolution MS (FAB, positive), calcd
for C107H184N2O22FPNa (M1Na)1: 1922.2960; found:
1922.2898.
2,6-Anhydro-3-deoxy-7-O-[2-deoxy-2-[(R)-3-(dodecanoyl-
oxy)tetradecanamido]-4-O-phosphono-3-O-[(R)-3-(tetra-
decanoyloxy)tetradecanoyl]-b-d-glucopyranosyl]-3-[(R)-
3-(hydroxy)tetradecanamido]-4-O-[(R)-3-(hydroxy)tetra-
decanoyl]-d-glycero-d-ido-heptonic acid (4a). To a solu-
tion of 24a (5.2 mg, 0.0027 mmol) in THF (1.0 ml), PtO2
(6.3 mg) was added, stirred for 18 h under hydrogen at
atmospheric pressure at 508C, and then ®ltered. The ®ltrate
was concentrated in vacuo, and separated by preparative
thin layer chromatography (CHCl3±EtOH±AcOH±H2O,
8:5:1:1). The part containing a product was eluted with
CHCl3±EtOH±AcOH±H2O (8:5:1:1) to give a mixture of
4a contaminated with silica gel. The mixture was dissolved
in CHCl3±MeOH±0.1 M HCl aq. (5:10:4) to remove the
silica gel, and extracted with CHCl3. The chloroform layer
was concentrated in vacuo to give 4a (4.2 mg, 88%) as a
powder. [a]2D6ˆ22.6 (cˆ0.18, CHCl3). IR: nmax (KBr)
C107H185N2O23PNa
1920.2963.
(M1Na)1:
1920.3003;
found:
2,6-Anhydro-3-deoxy-7-O-[2-deoxy-4-O-diphenylphos-
phono-2-[(R)-3-(dodecanoyloxy)tetradecanamido]-6-O-
methyl-3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-b-
d-glucopyranosyl]-3-[(R)-3-(hydroxy)tetradecanamido]-
4-O-[(R)-3-(hydroxy)tetradecanoyl]-d-glycero-d-ido-
heptonic acid (24b). Compound 23b (29.0 mg, 0.013
mmol) was treated as described in the formation of 24a
from 23a to give 24b (12.4 mg, 57%) as a powder. IR:
.
3351, 1734, 1662, 1468 cm21 1H NMR (400 MHz,
CD3OD±CDCl3 (10:1)) 5.32±5.12 (4H, m), 4.68 (1H, d,
Jˆ8.8 Hz), 4.50 (1H, d, Jˆ5.9 Hz), 4.33 (1H, dd, Jˆ10.2,
5.1 Hz), 4.21 (1H, q, Jˆ9.5 Hz), 4.08 (1H, d, Jˆ10.3 Hz),
4.02±3.95 (1H, m), 3.90±3.77 (6H, m), 3.65±3.57 (1H, m),
3.50±3.45 (1H, m), 2.68±2.65 (2H, m), 2.53±2.40 (4H, m),
2.37±2.26 (6H, m), 1.68±1.24 (120H, m), 0.90 (18H, t,
Jˆ6.6 Hz). High Resolution MS (FAB, negative), calcd
for C95H176N2O23P (M2H)2: 1744.2402; found: 1744.2352.
1
nmax (KBr) 3350 (broad), 1736 cm21. H NMR (400 MHz,
CD3OD) 7.41±7.36 (4H, m), 7.27±7.18 (6H, m), 5.44 (1H,
t, Jˆ9.5 Hz), 5.27±5.18 (2H, m), 4.95 (1H, d, Jˆ8.8 Hz),
4.71 (1H, q, Jˆ9.2 Hz), 4.27±4.22 (1H, m), 4.14 (1H, d,
Jˆ5.1 Hz), 4.04 (1H, d, Jˆ11.0 Hz), 4.02±3.94 (1H, m),
3.93±3.78 (3H, m), 3.77±3.68 (2H, m), 3.53±3.44 (3H,
m), 3.21 (3H, s), 2.61±2.40 (6H, m), 2.38±2.21 (4H, m),
2.13 (2H, t, Jˆ7.3 Hz), 1.68±1.17 (120H, m), 0.90 (18H, t,
Jˆ6.6 Hz). High Resolution MS (FAB, positive), calcd for
2,6-Anhydro-3-deoxy-7-O-[2-deoxy-2-[(R)-3-(dodecanoyl-
oxy)tetradecanamido]-6-O-methyl-4-O-phosphono-3-O-
[(R)-3-(tetradecanoyloxy)tetradecanoyl]-b-d-glucopyra-
nosyl]-3-[(R)-3-(hydroxy)tetradecanamido]-4-O-[(R)-3-
(hydroxy)tetradecanoyl]-d-glycero-d-ido-heptonic acid
(4b). Compound 24b (17.3 mg, 0.009 mmol) was treated
as described in the formation of 4a from 24a to give 4b
(7.8 mg, 49%) as a powder. [a]2D6ˆ24.0 (cˆ0.20, CHCl3).
C108H187N2O23PNa
(M1Na)1:
1934.3160;
found:
1934.3162. Anal. Calcd for C108H187N2O23P12H2O: C,
66.55; H, 9.88; N, 1.44; P, 1.59. Found: C, 66.22; H, 9.63;
N, 1.47; P, 1.62.
2,6-Anhydro-3-deoxy-7-O-[2,6-deoxy-4-O-diphenylphos-
phono-2-[(R)-3-(dodecanoyloxy)tetradecanamido]-6-
¯uoro-3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-b-
d-glucopyranosyl]-3-[(R)-3-(hydroxy)tetradecanamido]-
1
IR: nmax (KBr) 3351 (broad), 1734, 1664 cm21. H NMR
(400 MHz, CD3OD±CDCl3 (5:1)) 5.30±5.20 (4H, m), 4.67

T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
7701
(1H, d, Jˆ8.8 Hz), 4.48 (1H, d, Jˆ5.1 Hz), 4.37±4.33 (1H,
m), 4.14±4.05 (2H, m), 4.01±3.96 (1H, m), 3.92±3.87 (2H,
m), 3.82±3.76 (3H, m), 3.64±3.55 (3H, m), 3.42 (3H, s),
2.78±2.21 (12H, m), 1.67±1.23 (120H, m), 0.90 (18H, t,
Jˆ6.6 Hz). High Resolution MS (FAB, negative), calcd
for C96H178N2O23P (M2H)2: 1758.2558; found: 1758.2534.
decanoyloxy)tetradecanoyl]-b-d-glucopyranosyl]-3-
[(R)-3-(benzyloxy)tetradecanamido]-4-O-[(R)-3-(benzy-
loxy)tetradecanoyl]-3-deoxy-d-glycero-d-ido-heptonate
(25b). Compound 22b (23.3 mg, 0.012 mmol) was treated
as described in the formation of 25a from 22a to give 25b
(15.9 mg, 75%) as a powder. IR: nmax (KBr) 3339, 1742,
1
1654 cm21. H NMR (270 MHz, CDCl3) 7.39±7.13 (30H,
2,6-Anhydro-3-deoxy-7-O-[2,6-deoxy-2-[(R)-3-(dodecanoyl-
oxy)tetradecanamido]-6-¯uoro-4-O-phosphono-3-O-[(R)-3-
(tetradecanoyloxy)tetradecanoyl]-b-d-glucopyranosyl]-
3-[(R)-3-(hydroxy)tetradecanamido]-4-O-[(R)-3-(hydroxy)-
m), 6.82 (1H, s), 6.52 (1H, d, Jˆ9.0 Hz), 6.41 (1H, d,
Jˆ7.1 Hz), 5.46 (1H, dd, Jˆ10.0, 9.4 Hz), 5.30 (1H, dd,
Jˆ10.5, 9.3 Hz), 5.22 (1H, d, Jˆ8.2 Hz), 5.19±5.10 (1H,
m), 4.72±4.40 (6H, m), 4.36 (2H, d, Jˆ1.9 Hz), 4.05±3.58
(6H, m), 3.52±3.35 (4H, m), 3.21 (3H, s), 2.65±2.14 (8H,
m), 1.87 (3H, s), 1.78±1.11 (82H, m), 0.88 (12H, t, Jˆ
6.4 Hz). High Resolution MS (FAB, positive), calcd for
tetradecanoyl]-d-glycero-d-ido-heptonic
acid
(4c).
Compound 24c (7.7 mg, 0.0040 mmol) was treated as
described in the formation of 4a from 24a to give 4c
(5.2 mg, 73%) as a powder. [a]2D5ˆ23.7 (cˆ0.20, CHCl3).
C111H163N2O21PNa
(M1Na)1:
1914.1384;
found:
1
IR: nmax (KBr) 3362, 1734, 1662, 1467 cm21. H NMR
1914.1450. Anal. Calcd for C111H163N2O21P: C, 70.45; H,
8.68; N, 1.48; P, 1.64. Found: C, 70.50; H, 8.86; N, 1.45; P,
1.75.
(400 MHz, CD3OD±CDCl3 (10:1)) 5.32±5.24 (4H, m),
4.68 (1H, d, Jˆ9.5 Hz), 4.50 (1H, d, Jˆ5.9 Hz), 4.33 (1H,
dd, Jˆ10.6, 6.2 Hz), 4.22 (1H, q, Jˆ8.8 Hz), 4.07 (1H, d,
Jˆ9.5 Hz), 4.00±3.97 (1H, m), 3.90±3.85 (1H, m), 3.82±
3.62 (6H, m), 3.60±3.48 (2H, m, including 1H, t, Jˆ8.8 Hz
at 3.60 ppm), 2.72±2.62 (2H, m), 2.55±2.40 (4H, m), 2.37±
2.23 (6H, m), 1.67±1.25 (120H, m), 0.90 (18H, t, Jˆ
6.6 Hz). High Resolution MS (FAB, negative), calcd for
C95H175N2O22FP (M2H)2: 1746.2358; found: 1746.2345.
Diphenylmethyl 2,6-anhydro-7-O-[2-acetamido-2,6-deoxy-
4-O-diphenylphosphono-6-¯uoro-3-O-[(R)-3-(tetradecanoyl-
oxy)tetradecanoyl]-b-d-glucopyranosyl]-3-[(R)-3-
(benzyloxy)tetradecanamido]-4-O-[(R)-3-(benzyloxy)-
tetradecanoyl]-3-deoxy-d-glycero-d-ido-heptonate (25c).
Compound 22c (122 mg, 0.061 mmol) was treated as
described in the formation of 25a from 22a to give 25c
(65.1 mg, 57%) as a powder. IR: nmax (KBr) 3356, 1733,
1654, 1533, 1193 cm21. 1H NMR (400 MHz, CDCl3) 7.36±
7.08 (30H, m), 6.82 (1H, s), 6.55 (1H, d, Jˆ7.3 Hz), 6.52
(1H, d, Jˆ9.5 Hz), 5.53 (1H, t, Jˆ9.5 Hz), 5.38±5.28 (2H,
m), 5.15±5.13 (1H, m), 4.67±4.60 (2H, m), 4.57±4.45 (4H,
m), 4.39±4.32 (2H, m), 4.07±4.04 (1H, m), 3.86±3.83 (1H,
m), 3.73±3.56 (5H, m), 3.40±3.33 (2H, m), 2.61 (1H, dd,
Jˆ7.3, 15.4 Hz), 2.41 (2H, dd, Jˆ5.1, 15.4 Hz), 2.25±2.11
(5H, m), 1.87 (3H, s), 1.66±1.25 (82H, m), 0.88 (12H, t,
Jˆ6.6 Hz). High Resolution MS (FAB, positive), calcd for
C110H160N2O20FPNa (M1Na)1: 1902.1184; found:
1902.1167. Anal. Calcd for C110H160N2O20FP: C, 70.26;
H, 8.58; N, 1.49; F, 1.01; P, 1.65. Found: C, 69.98; H,
8.08; N, 1.34; F, 1.12; P, 1.77.
Diphenylmethyl 2,6-anhydro-7-O-[2-acetamido-6-O-benzyl-
oxycarbonyl-2-deoxy-4-O-diphenylphosphono-3-O-[(R)-3-
(tetradecanoyloxy)tetradecanoyl]-b-d-glucopyranosyl]-3-
[(R)-3-(benzyloxy)tetradecanamido]-4-O-[(R)-3-(benzyl-
oxy)tetradecanoyl]-3-deoxy-d-glycero-d-ido-heptonate
(25a). To a solution of 22a (99.9 mg, 0.046 mmol) in acetic
acid (2.0 ml), zinc dust (310 mg, 4.74 mmol) was added.
After vigorously stirring for 5 h at room temperature, the
solution was ®ltered through Celite to remove the zinc dust
and concentrated in vacuo to give a crude product. To a
solution of this crude product in CH2Cl2 (2.0 ml), acetic
acid (12 mg) in CH2Cl2 (2.0 ml) and EDCI´HCl (17.8 mg,
0.093 mmol) were added at room temperature, and the
mixture was allowed to stand for 4 h at room temperature.
Then, the reaction mixture was concentrated in vacuo, and
chromatographed on a silica gel column eluted with hexane±
EtOAc (2:1) to give 25a (43.5 mg, 47%) as a viscous oil. IR:
2,6-Anhydro-7-O-[2-acetamido-2-deoxy-4-O-diphenyl-
phosphono-3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-
b-d-glucopyranosyl]-3-deoxy-3-[(R)-3-(hydroxy)tetra-
decanamido]-4-O-[(R)-3-(hydroxy)tetradecanoyl]-d-
glycero-d-ido-heptonic acid (26a). To a solution of 25a
(15.1 mg, 0.008 mmol) in ethanol (1.0 ml), 20% Pd(OH)2
on carbon (16.0 mg) was added, and the mixture was stirred
for 7 h under hydrogen at atmospheric pressure at room
temperature, and ®ltered through Celite. The ®ltrate was
concentrated in vacuo, and separated by preparative thin
layer chromatography (CHCl3±MeOH, 5:1). The part
containing a product was eluted with CHCl3±MeOH (5:1)
to give a mixture of 26a contaminated with silica gel. The
mixture was dissolved in CHCl3±MeOH±0.1 M HCl aq.
(5:10:4) to remove the silica gel, and extracted with
CHCl3. The chloroform layer was concentrated in vacuo
to give 26a (7.8 mg, 63%) as a powder. IR: nmax (KBr)
1
nmax (KBr) 3360, 1740, 1652, 1531, 1192 cm21. H NMR
(500 MHz, CDCl3) 7.35±7.11 (35H, m), 6.82 (1H, s), 6.53
(1H, d, Jˆ3.9 Hz), 6.52 (1H, d, Jˆ6.8 Hz), 5.50 (1H, t,
Jˆ9.3 Hz), 5.36±5.30 (2H, m), 5.18±5.08 (1H, m), 5.12
(1H, AB±q, Jˆ12.2 Hz), 5.03 (1H, AB-q, Jˆ12.2 Hz),
4.68±4.62 (2H, m, including 1H, AB-q, Jˆ5.9 Hz at
4.62 ppm), 4.54 (1H, AB-q, Jˆ11.7 Hz), 4.49±4.45 (2H, m,
including 1H, AB-q, Jˆ10.7 Hz at 4.47 ppm), 4.40±4.33
(3H, m), 4.18 (1H, dd, Jˆ12.2, 4.4 Hz), 4.03 (2H, d,
Jˆ9.8 Hz), 3.88±3.81 (1H, m), 3.78±3.73 (1H, m), 3.68±
3.55 (4H, m), 3.38±3.31 (2H, m), 2.61 (1H, dd, Jˆ15.4,
7.3 Hz), 2.46±2.38 (2H, m), 2.28±2.11 (5H, m), 1.87 (3H,
s), 1.75±1.08 (85H, m), 0.88 (12H, t, Jˆ6.9 Hz). High Reso-
lution MS (FAB, positive), calcd for C118H167N2O23PNa
(M1Na)1: 2034.1595; found: 2034.1606. Anal. Calcd for
C118H167N2O23P: C, 70.42; H, 8.36; N, 1.39; P, 1.54.
Found: C, 70.14; H, 8.22; N, 1.28; P, 1.27.
1
3373, 1738, 1662, 1490, 1189 cm21. H NMR (400 MHz,
CD3OD) 7.41±7.36 (4H, m), 7.26±7.18 (6H, m), 5.41 (1H,
t, Jˆ9.9 Hz), 5.30 (1H, t, Jˆ10.3 Hz), 5.11 (1H, t,
Jˆ6.2 Hz), 4.76 (1H, d, Jˆ8.8 Hz), 4.71 (1H, q,
Jˆ9.2 Hz), 4.49±4.46 (1H, m), 4.36±4.28 (1H, m), 4.11
Diphenylmethyl 2,6-anhydro-7-O-[2-acetamido-2-deoxy-
4-O-diphenylphosphono-6-O-methyl-3-O-[(R)-3-(tetra-

7702
T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
(1H, d, Jˆ11.0 Hz), 3.97±3.75 (6H, m), 3.67±3.58 (2H, m),
3.53 (1H, t, Jˆ8.8 Hz), 2.54±2.39 (4H, m), 2.32±2.21 (2H,
m), 2.16 (2H, t, Jˆ7.3 Hz), 1.94 (3H, s), 1.53±1.18 (82H,
m), 0.91±0.88 (12H, m). High Resolution MS (FAB, posi-
tive), calcd for C83H139N2O21PNa (M1Na)1: 1553.9506;
found: 1553.9490. Anal. Calcd for C83H139N2O21P12H2O:
C, 63.58; H, 9.19; N, 1.79; P, 1.98. Found: C, 63.21; H, 8.72;
N, 1.77; P, 2.02.
2,6-Anhydro-7-O-[2-acetamido-2-deoxy-4-O-phosphono-
3-O-[(R)-3-(tetradecanoyloxy)tetradecanoyl]-b-d-gluco-
pyranosyl]-3-deoxy-3-[(R)-3-(hydroxy)tetradecana-
mido]-4-O-[(R)-3-(hydroxy)tetradecanoyl]-d-glycero-d-
ido-heptonic acid (5a). To a solution of 26a (6.6 mg,
0.004 mmol) in THF (1.0 ml), PtO2 (8.4 mg) was added,
and the mixture was stirred for 20 h under hydrogen at
atmospheric pressure at room temperature, and then ®ltered.
The ®ltrate was concentrated in vacuo, and separated by
preparative thin layer chromatography (CHCl3±EtOH±
AcOH±H2O, 8:5:1:1). The part containing a product was
eluted with CHCl3-EtOH-AcOH-H2O (8:5:1:1) to give a
mixture of 5a contaminated with silica gel. The mixture
was dissolved in CHCl3±MeOH±0.1 M HCl aq. (5:10:4)
to remove the silica gel, and extracted with CHCl3. The
chloroform layer was concentrated in vacuo to give 5a
(4.9 mg, 82%) as a powder. [a]2D5ˆ27.8 (cˆ0.20, CHCl3).
2,6-Anhydro-7-O-[2-acetamido-2-deoxy-4-O-diphenyl-
phosphono-6-O-methyl-3-O-[(R)-3-(tetradecanoyloxy)-
tetradecanoyl]-b-d-glucopyranosyl]-3-deoxy-3-[(R)-3-
(hydroxy)tetradecanamido]-4-O-[(R)-3-(hydroxy)tetra-
decanoyl]-d-glycero-d-ido-heptonic acid (26b). To a solu-
tion of 25b (14.6 mg, 0.008 mmol) in EtOAc (3.0 ml), 20%
Pd(OH)2 on carbon (15.2 mg) was added, and the mixture
was stirred for 16 h under hydrogen at atmospheric pressure
at room temperature, and then, ®ltered through Celite. The
®ltrate was concentrated in vacuo, and separated by prepara-
tive thin layer chromatography (CHCl3±MeOH, 8:1). The
part containing a product was eluted with CHCl3±MeOH
(5:1) to give a mixture of 26b contaminated with silica gel.
The mixture was dissolved in CHCl3±MeOH±0.1 M HCl
aq. (5:10:4) to remove the silica gel, and extracted with
CHCl3. The chloroform layer was concentrated in vacuo
to give 26b (7.7 mg, 63%) as a powder. IR: nmax (KBr)
1
IR: nmax (KBr) 3368, 1737, 1664, 1466 cm21. H NMR
(500 MHz, CD3OD±CDCl3 (10:1)) 5.26±5.12 (3H, m),
4.62 (1H, d, Jˆ8.8 Hz), 4.31±4.18 (3H, m), 4.09 (1H, d,
Jˆ9.5 Hz), 4.03±3.87 (4H, m), 3.82±3.73 (3H, m), 3.50±
3.38 (2H, m), 2.74±2.60 (2H, m), 2.52±2.38 (2H, m), 2.36±
2.22 (4H, m), 1.98 (3H, s), 1.63±1.58 (4H, m), 1.45±1.41
(6H, m), 1.38±1.23 (72H, m), 0.90 (12H, t, Jˆ7.3±6.6 Hz).
High Resolution MS (FAB, negative), calcd for
C71H130N2O21P (M2H)2: 1377.8904; found: 1377.8844.
Anal. Calcd for C71H131N2O21P1H2O: C, 61.01; H, 9.59;
N, 2.00; P, 2.22. Found: C, 61.24; H, 9.79; N, 2.14; P, 1.94.
3500±3300 (broad), 1740, 1655 cm21 1H NMR (400
.
MHz, CD3OD) 8.16 (1H, d, Jˆ10.0 Hz), 7.41±7.37 (4H,
m), 7.27±7.19 (6H, m), 5.41 (1H, t, Jˆ9.9 Hz), 5.26 (1H, t,
Jˆ9.5 Hz), 5.10 (1H, t, Jˆ5.9 Hz), 4.97 (1H, d, Jˆ8.1 Hz),
4.72 (1H, q, Jˆ9.2 Hz), 4.27±4.23 (1H, m), 4.16 (1H, d,
Jˆ5.1 Hz), 4.04 (1H, d, Jˆ12.5 Hz), 3.97±3.82 (4H, m),
3.77±3.70 (2H, m), 3.54±3.41 (3H, m), 3.22 (3H, s), 2.50±
2.37 (4H, m), 2.34±2.21 (2H, m), 2.15 (2H, t, Jˆ7.3 Hz), 2.00
(3H, s), 1.55±1.25 (82H, m), 0.90 (12H, t, Jˆ7.3±6.6 Hz).
High Resolution MS (FAB, positive), calcd for
C84H141N2O21PNa (M1Na)1: 1567.9662; found: 1567.9694.
Anal. Calcd for C84H141N2O21P: C, 65.26; H, 9.19; N, 1.81; P,
2.00. Found: C, 64.89; H, 9.04; N, 1.64; P, 1.97.
2,6-Anhydro-7-O-[2-acetamido-2-deoxy-6-O-methyl-4-
O-phosphono-3-O-[(R)-3-(tetradecanoyloxy)tetradeca-
noyl]-b-d-glucopyranosyl]-3-deoxy-3-[(R)-3-(hydroxy)-
tetradecanamido]-4-O-[(R)-3-(hydroxy)tetradecanoyl]-
d-glycero-d-ido-heptonic acid (5b). Compound 26b
(7.0 mg, 0.0045 mmol) was treated as described in the
formation of 5a from 26a to give 5b (6.8 mg, 99%) as a
powder. [a]2D6ˆ26.6 (cˆ0.20, CHCl3). IR: nmax (KBr)
3362, 1737, 1665 cm21 1H NMR (400 MHz, CD3OD)
.
5.30 (1H, dd, Jˆ10.7, 8.8 Hz), 5.23±5.15 (2H, m), 4.66
(1H, d, Jˆ8.5 Hz), 4.50 (1H, d, Jˆ5.7 Hz), 4.34±4.29
(1H, m), 4.22±4.19 (1H, m), 4.10 (1H, d, Jˆ1.4 Hz),
4.00±3.94 (1H, m), 3.88±3.73 (5H, m), 3.62±3.57 (2H,
m), 3.54 (1H, t, Jˆ9.1 Hz), 3.41 (3H, s), 2.72±2.22 (8H,
m), 1.94 (3H, s), 1.62±1.56 (4H, m), 1.48±1.40 (6H, m),
1.39±1.23 (72H, m), 0.91 (12H, t, Jˆ7.0±6.6 Hz). High
Resolution MS (FAB, negative), calcd for C72H132N2O21P
(M2H)2: 1391.9060; found: 1391.9093. Anal. Calcd for
C72H133N2O21P1H2O: C, 61.25; H, 9.64; N, 1.98; P, 2.19.
Found: C, 61.53; H, 9.50; N, 1.99; P, 2.23.
2,6-Anhydro-7-O-[2-acetamido-2,6-deoxy-4-O-diphenyl-
phosphono-6-¯uoro-3-O-[(R)-3-(tetradecanoyloxy)tetra-
decanoyl]-b-d-glucopyranosyl]-3-deoxy-3-[(R)-3-
(hydroxy)tetradecanamido]-4-O-[(R)-3-(hydroxy)tetra-
decanoyl]-d-glycero-d-ido-heptonic acid (26c). Com-
pound 25c (54.5 mg, 0.029 mmol) was treated as
described in the formation of 26a from 25a to give 26c
(29.1 mg, 65%) as an amorphous ®lm. IR: nmax (KBr)
1
3345, 1738, 1662, 1491, 1189 cm21. H NMR (400 MHz,
CD3OD) 7.41±7.37 (4H, m), 7.27±7.17 (6H, m), 5.46 (1H,
t, Jˆ9.9 Hz), 5.27 (1H, t, Jˆ9.5 Hz), 5.13±5.04 (1H, m),
4.81 (1H, d?, unclear due to overlapping with solvent's
peak), 4.70 (1H, q, Jˆ9.3 Hz), 4.60 (1H, d, Jˆ11.0 Hz),
4.49 (1H, d, Jˆ11.0 Hz), 4.40±4.36 (1H, m), 4.27±4.23
(1H, m), 4.18±4.14 (1H, m), 4.04 (1H, d, Jˆ12.5 Hz),
3.99±3.81 (4H, m), 3.76±3.68 (1H, m), 3.42 (3H, t,
Jˆ9.5 Hz), 2.50±2.41 (4H, m), 2.34±2.21 (2H, m), 2.16
(2H, t, Jˆ7.3 Hz), 2.01 (3H, s), 1.55±1.20 (82H, m), 0.90
(12H, t, Jˆ6.6 Hz). High Resolution MS (FAB, positive),
calcd for C83H138N2O20FPNa (M1Na)1: 1555.9462; found:
1555.9437. Anal. Calcd for C83H138N2O20FP12H2O: C,
63.50; H, 9.12; N, 1.78; F, 1.21; P, 1.97. Found: C, 63.37;
H, 8.66; N, 1.76; F, 1.41; P, 2.11.
2,6-Anhydro-7-O-[2-acetamido-2,6-deoxy-6-¯uoro-4-O-
phosphono-3-O-[(R)-3-(tetradecanoyloxy)tetradeca-
noyl]-b-d-glucopyranosyl]-3-deoxy-3-[(R)-3-(hydroxy)-
tetradecanamido]-4-O-[(R)-3-(hydroxy)tetradecanoyl]-
d-glycero-d-ido-heptonic acid (5c). Compound 26c
(17.1 mg, 0.0111 mmol) was treated as described in the
formation of 5a from 26a to give 5c (13.3 mg, 87%) as a
powder. [a]2D6ˆ218.6 (cˆ0.20, CHCl3). IR: nmax (KBr)
3363, 1734, 1662, 1466 cm21
.
1H NMR (400 MHz,
CD3OD±CDCl3(10:1)) 5.23±5.14 (3H, m), 4.72±4.57 (2H,
m), 4.33±4.28 (2H, m), 4.18±4.12 (2H, m), 4.00±3.88 (3H,
m), 3.78±3.56 (4H, m), 3.32 (1H, t, Jˆ1.5 Hz), 2.74 (1H, d,
Jˆ9.5, 6.6 Hz), 2.63 (1H, dd, Jˆ9.5, 6.6 Hz), 2.52±2.40

T. Mochizuki et al. / Tetrahedron 56 (2000) 7691±7703
7703
3. (a) Imoto, M.; Yoshimura, H.; Sakaguchi, N.; Kusumoto, S.;
Shiba, T. Tetrahedron Lett. 1985, 26, 1545±1548. (b) Imoto, M.;
Yoshimura, H.; Shimamoto, T.; Sakaguchi, N.; Kusumoto, S.;
Shiba, T. Bull. Chem. Soc. Jpn 1987, 60, 2205±2214.
(c) Takahashi, T.; Nakamoto, S.; Ikeda, K.; Achiwa, K.
Tetrahedron Lett. 1986, 27, 1819±1822.
(2H, m), 2.36±2.22 (4H, m), 1.99 (3H, s), 1.63±1.58 (4H,
m), 1.50±1.42 (6H, m), 1.38±1.22 (72H, m), 0.89 (12H, t,
Jˆ7.3±6.6 Hz). High Resolution MS (FAB, negative),
calcd for C71H129N2O20FP (M2H)2: 1379.8860; found:
1379.8854. Anal. Calcd for C71H130N2O20FP: C, 61.72; H,
9.48; N, 2.03; F, 1.38; P, 2.24. Found: C, 61.55; H, 9.36; N,
1.76; F, 1.19; P, 2.11.
4. Galanos, C. Z. Immunitatsforsch. Exp. Klin. Immunol. 1975,
149, 214±229.
Method for biological activity measurement12
5. Rietschel, E. T.; Kirikae, T.; Schade, F. U.; Ulmer, A. J.; Holst,
O.; Brade, H.; Schmidt, G.; Mamat, U.; Grimmecke, H-D.;
È
Kusumoto, S.; Zahringer, U. Immunobiology 1993, 187, 169±190.
The sources of the materials used in the study are as follows:
lipopolysaccharide (LPS) from E. coli serotype 026:B618
and 12-O-tetradecanoylphorbor acetate (TPA) were from
SIGMA, St. Louis, MO; RPMI-1640 medium, fetal bovine
serum (FBS), and newborn calf serum (NBCS) were from
GIBCO, Grand Island, NY; and human tumor necrosis
factor-a enzyme-linked immunosorbent assay (TNFa
ELISA) kit was from Genzyme, Cambridge, MA.
6. (a) Qureshi, N.; Honovich, J. P.; Hara, H.; Cotter, R. J.;
Takayama, K. J. Biol. Chem. 1988, 263, 5502±5504. (b) Qureshi,
N.; Takayama, K.; Kurtz, R. Infect. Immun. 1991, 59, 441±444.
7. (a) Christ, W. J.; McGuinness, P. D.; Asano, O.; Wang, Y.;
Mullarkey, M. A.; Perez, M.; Hawkins, L. D.; Blythe, T. A.;
Dubuc, G. R.; Robidoux, A. L. J. Am. Chem. Soc. 1994, 116,
3637±3638. (b) Christ, W. J.; Asano, O.; Robidoux, A. L. C.;
Perez, M.; Wang, Y.; Dubuc, G. R.; Gavin, W. E.; Hawkins,
L. D.; McGuinness, P. D.; Mullarkey, M. A.; Lewis, M. D.;
Kishi, Y.; Kawata, T.; Bristol, J. R.; Rose, J. R.; Rossignol, D. P.;
Kobayashi, S.; Hishinuma, I.; Kimura, A.; Asakawa, N.;
Katayama, K.; Yamatsu, I. Science 1995, 268, 80±83.
8. Kusama, T.; Soga, T.; Ono, Y.; Kumazawa, E.; Shioya, E.;
Nakayama, K.; Uoto, K.; Osada, Y. Chem. Pharm. Bull. 1991,
39, 3244±3253.
Cell culture: Human monoblastic U937 cells were main-
tained in RPMI-1640 medium supplemented with 10%
FBS, 100 units/ml of penicillin and 100 mg/ml of strepto-
mycin (growth medium).
Production of TNFa by U937 cells: U937 cells (1£104/
200ml/well) were plated in 96-well plates (Corning,
Cambridge, MA), and were cultured in the presence of
TPA (30 ng/ml) for 72 h at 378C. After removing the super-
natant, the cells were incubated in 200 ml of fresh RPMI-
1640 medium containing 10% NBCS, in the absence or the
presence of 30 ng/ml of LPS with graded concentrations of
the compounds in a humidi®ed atmosphere of 5% CO2 for
4.5 h at 378C. After incubation, the amount of TNFa
produced in the culture supernatants was determined using
the TNFa ELISA kits. As a control, the amount of TNFa
produced by the U937 cells, which were stimulated with
30 ng/ml of LPS in the absence of compounds, was used.
The concentrations (nM) of compounds required to inhibit
the LPS-induced TNFa production by U937 cells by 50%
(IC50) was calculated from the control amount. All experi-
ments were carried out at least twice, showing the data are
reproducible. The representative data are shown in Figs. 3
and 4.
9. Liu, W.-C.; Oikawa, M.; Fukase, K.; Suda, Y.; Kusumoto, S.
Bull. Chem. Soc. Jpn 1999, 72, 1377±1385.
10. Wang, M.-H.; Flad, H-D.; Feist, W.; Musehold, J.; Kusumoto,
S.; Brade, H.; Gerdes, J.; Rietschel, H. T.; Ulmer, A. J. Lymphokine
Cytokine Res. 1992, 11, 23±31.
11. (a) Matsuura, M.; Kojima, Y.; Homma, J. Y.; Kubota, Y.;
Yamamoto, A.; Kiso, M.; Hasegawa, A. FEBS Lett. 1984, 167,
226±230. (b) Kiso, M.; Ogawa, Y.; Fujishima, Y.; Fujita, M.;
Tanaka, S.; Hasegawa, A. J. Carbohydr. Chem. 1987, 6, 625±638.
12. Shiozaki, M.; Kurakata, S.; Tatsuta, T.; Maeda, H.; Nishijima,
M. Tetrahedron 1997, 53, 16041±16060.
13. Mochizuki, T.; Iwano, Y.; Shiozaki, M.; Kurakata, S.; Kanai,
S.; Nishijima, M. Carbohydr. Res. 2000, 324, 225±230.
14. Shiozaki, M.; Deguchi, N.; Macindoe, W. M.; Arai, M.;
Miyazaki, H.; Mochizuki, T.; Tatsuta, T.; Ogawa, J.; Maeda, H.;
Kurakata, S. Carbohydr. Res. 1996, 283, 27±51.
15. (a) Haines, L. M.; Singleton, E. J. Chem. Soc., Dalton Trans.
1972, 1891±1896. (b) Oltvoort, J. J.; van Boeckel, C. A. A.; de
Koning, J. H.; van Boom, J. H. Synthesis 1981, 305±308.
16. Schmidt, R. R. Angew. Chem. Int. Ed. Engl. 1986, 25, 212±
235.
Acknowledgements
This study was supported, in part, by the Social Insurance
Agency Fund commissioned by the Japan Health Sciences
Foundation.
17. Shiozaki, M.; Arai, M.; Macindoe, W. M.; Mochizuki, T.;
Wakabayashi, T.; Kurakata, S.; Tatsuta, T.; Maeda, H.; Nishijima,
M. Bull. Chem. Soc. Jpn 1997, 70, 1149±1161.
18. (a) Dickerson, R.; Manzo, C.; Charland, S.; Settle, R.; Stein,
T.; Kuhl, D.; Rajter, J. J. Surg. Res. 1995, 58, 260±266. (b) Michel,
O.; Duchateau, J.; Plat, G.; Cantinieaux, B.; Hotimsky, A.; Gerain,
J.; Sergysels, R. Clin. Exp. Allergy 1995, 25, 73±79.
References
1. Westphal, O.; LuÈderitz, O. Angew. Chem. 1954, 66, 407±417.
2. Galanos, C.; LuÈderitz, O.; Rietschel, E. T.; Westphal, O. Int.
Rev. Biochem. 1977, 14, 239±335.