Novel design and synthesis of a radioiodinated glycolipid analog as an acceptor substrate for N-acetylglucosaminyltransferase v

Article abstract of DOI:10.1002/jlcr.3063

Guided by the known molecular recognition interactions between N-acetylglucosaminyltransferase V (GnT-V) and certain synthetic substrates, we synthesized a radiolabeled double-stranded glycolipid composed of a long-chain alkyl unit and a radioiodinated phenylalkyl unit, [¹²⁵I]-2-[N-(2- hydroxy-3-hexadecyloxy)propyl-15-(4-iodophenyl)pentadecanecarboxamido]ethyl 2-acetamido-2-deoxy-β-d-glucopyranosyl-(1→2)-α-d-mannopyranosyl- (1→6)-β-d-glucopyranoside ([¹²⁵I]2), as a novel intravital glycolipid mimic substrate of GnT-V. The radioactive iodine (¹²⁵I) was incorporated via iododestannylation of the phenyltributyltin derivative, 2-[N-(2-acetoxy-3-hexadecyloxy)propyl-15-(4-tributylstannylphenyl) pentadecanecarboxamido]ethyl 3,4,6-tri-O-acetyl-2-acetamido-2-deoxy-β-d- glucopyranosyl-(1→2)-3,4,6-O-acetyl-α-d-mannopyranosyl-(1→6)-2, 3,4-tri-O-acetyl-β-d-glucopyranoside (26). Subsequent deacetylation at the final step afforded [¹²⁵I]2. Copyright

Full text of DOI:10.1002/jlcr.3063

Research Article
Received 6 February 2013,
Revised 22 April 2013,
Accepted 23 April 2013
Published online 4 September 2013 in Wiley Online Library
(wileyonlinelibrary.com) DOI: 10.1002/jlcr.3063
Novel design and synthesis of a
radioiodinated glycolipid analog as an
acceptor substrate for
N-acetylglucosaminyltransferase V
a
Kenji Arimitsu,a,b Hiroyuki Kimura,a Tetsuya Kajimoto,b,c Masahiro Ono,
*
Yoshiro Ohmomo,d Masayuki Yamashita,b Manabu Node,b
a
*
and Hideo Saji
Guided by the known molecular recognition interactions between N-acetylglucosaminyltransferase V (GnT-V) and
certain synthetic substrates, we synthesized a radiolabeled double-stranded glycolipid composed of a long-chain
alkyl unit and a radioiodinated phenylalkyl unit, [125I]-2-[N-(2-hydroxy-3-hexadecyloxy)propyl-15-(4-iodophenyl)
pentadecanecarboxamido]ethyl 2-acetamido-2-deoxy-b-D-glucopyranosyl-(1!2)-a-D-mannopyranosyl-(1!6)-b-D-glucopyranoside
([125I]2), as a novel intravital glycolipid mimic substrate of GnT-V. The radioactive iodine (125I) was incorporated via
iododestannylation of the phenyltributyltin derivative, 2-[N-(2-acetoxy-3-hexadecyloxy)propyl-15-(4-tributylstannylphenyl)
pentadecanecarboxamido]ethyl 3,4,6-tri-O-acetyl-2-acetamido-2-deoxy-b-D-glucopyranosyl-(1!2)-3,4,6-O-acetyl-a-D-
mannopyranosyl-(1!6)-2,3,4-tri-O-acetyl-b-D-glucopyranoside (26). Subsequent deacetylation at the final step afforded
[
125I]2.
Keywords: radioiodinated glycolipid analog; N-acetylglucosaminyltransferase V (GnT-V); trisaccharide; double-stranded lipid
ꢁ1.69.† Increasing the hydrophobicity of 1a is expected to be
Introduction
important for increasing the molecule’s permeation into the Golgi
Glycoconjugates on cell surfaces play critical roles in
apparatus in the design of reliable and useful substrates. We next
sought to synthesize the trisaccharide derivatives [125I]1b and [125I]
biological events, such as development, carcinogenesis, and
malignant transformation. Recent studies revealed a close
relationship between the production of unusual sugar chains
on cell surfaces and malignant cell transformation; a variety
1c, which bear longer alkyl chains than the p-iodophenylethyl group,
for example, the p-iodophenyloctyl or p-iodophenylpentadecyl
groups (Figure 1).12 125I]1b and [125I]1c were expected to display a
[
of highly branched oligosaccharides have been identified on
tumor cell surfaces. Several studies over the past two decades
have demonstrated that N-acetylglucosaminyltransferase
V (GnT-V) is one of the most relevant glycosyltransferases
for tumor invasion and metastasis. Since Cummings and his
colleagues reported GnT-V expression in mouse lymphoma
cells in 1982,1 GnT-V has been shown to be highly expressed
in a variety of cancer cell lines starting from the early stages
of tumorigenesis.2–8 Some biological studies of GnT-V-
deficient mice have directly demonstrated the essential role
of the transferase in the growth and metastasis of tumor
cells.9,10
higher permeability through the cell membranes and into the Golgi
aDepartment of Patho-functional Bioanalysis, Graduate School of Pharmaceutical
Sciences, Kyoto University, 46-29 Yoshida Shimoadachi-cho, Sakyo-ku, Kyoto, 606-
8501, Japan
bDepartment of Pharmaceutical Manufacturing Chemistry, Kyoto Pharmaceutical
University, 1 Shichono-cho, Yamashina-ku, Kyoto, 607-8414, Japan
cRitsumeikan University, 1-1-1 Nojihigashi, Kusatsu, Shiga, 525-8577, Japan
dOsaka University of Pharmaceutical Sciences, 4-1-20 Nasahara, Takatsuki,
Osaka, 569-1094, Japan
With the aim of developing an imaging tracer for GnT-V activity,
we previously reported the synthesis of the radioiodinated
trisaccharide derivative, 2-(4-[125I]iodophenyl)ethyl 2-deoxy-2-
phthalimido-b-D-glucopyranosyl-(1!2)-a-D-mannopyranosyl-(1!6)-
b-D-glucopyranoside ([125I]1a, [125I]IPGMG), which has a Km of
23.7 mM and a Vmax of 159 pmol/(hꢀmg) toward GnT-V in vitro
*Correspondence to: Hideo Saji, Department of Patho-functional Bioanalysis,
Graduate School of Pharmaceutical Sciences, Kyoto University, 46-29 Yoshida
Shimoadachi-cho, Sakyo-ku, Kyoto 606-8501, Japan.
E-mail: hsaji@pharm.kyoto-u.ac.jp
Correspondence to: Tetsuya Kajimoto, Ritsumeikan University, 1-1-1 Nojihigashi,
Kusatsu, Shiga, 525-8577, Japan.
(Figure 1).11 125I]1a was not expected to incorporate into the Golgi
[
E-mail: kajimoto@fc.ritsumei.ac.jp
† Calculated logP values were obtained computationally using ACD/Labs
v.12.0 (Advanced Chemistry Development, Toronto, Ontario, Canada).
apparatus, where GnT-V is excessively expressed, because of a low
hydrophobicity; the calculated value of log P (clogP) for [125I]1a was
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K. Arimitsu et al.
Figure 1. Structures of the 125I-labeled trisaccharide derivatives [125I]1a–c and [125I]2.
apparatus than [125I]1a based on the higher clogP values ([125I]1b (10) afforded 7. The o-nitrobenzyl group was chosen as the
clogP = 1.14; [125I]1c clogP = 4.86).† In fact, [125I]1c showed better protecting group on the aglycon glucoside moiety 10 because it
uptake profile in comparison with [125I]1a and [125I]1b ([125I]1c could be selectively cleaved by sunlight photolysis16,17 after the
[
125I]1b> [125I]1a).{
construction of the trisaccharyl moiety.
Certain intravital glycolipids, such as sphingoglycolipids or
First, we synthesized the glucoside 10 as a glycosyl acceptor.
glyceroglycolipids, consist of double-strand structures with two The glycosylation of 2-(2-nitrobenzyloxy) ethanol (11)18 with
long alkyl chains. The structures of these glycolipids could play penta-O-acetyl-b-D-glucopyranose in the presence of BF3ꢀEt2O
an important role in their interactions with other glycolipids gave the glucoside 12, which was then deacetylated. The free
or cells. Here, we designed an alternative GnT-V-specific b-D-glucoside 13 obtained was treated with trityl chloride to
radioiodinated glycolipid analog that could be incorporated into afford the 6-O-tritylated glucoside 14. Subsequent treatment of
the cell. A neoglycolipid aglycon moiety was designed to have a 14 with benzyloxy-2,2,2-trichloroacetimidate in the presence of
double-strand structure composed of
a long alkyl chain a catalytic amount of trimethylsilyl trifluoromethanesulfonate
and a radioiodinated phenylalkyl unit. We synthesized a novel afforded the tribenzyl ether 15. Treatment of 15 with hydrogen
125I-labeled glycolipid [125I]2 via a key intermediate, from which chloride in 1,4-dioxane, to cleave the trityl group, gave 10
the aglycon moiety could be facilely modified in a few steps just (Scheme 2).
prior to incorporating the radioiodine atom (Figure 1).
Next, the trisaccharide 7 was synthesized from the glucoside
10 by repeated glycosylation and further transformation of the
functional group using the route depicted in Scheme 3.13,14
Glycosylation of acceptor 10 with the glycosyl donor 9 in the
presence of N-iodosuccinimide and silver trifluoromethanesulfonate
in dichloromethane yielded (2-nitrobenzyloxy)ethyl 2-O-acetyl-3-O-
benzyl-4,6-O-benzylidene-a-D-mannopyranosyl-(1!6)-2,3,4-
tri-O-benzyl-b-D-glucopyranoside (16) (84%). Conventional
deacetylation of 16 gave the disaccharide 17, which was used as a
glycosyl acceptor (91%), in a subsequent glycosylation step, using 8
as the glycosyl donor, to afford 2-(2-nitrobenzyloxy)ethyl 3,4,6-tri-O-
acetyl-2-deoxy-2-phthalimido-b-D-glucopyranosyl-(1!2)-3-O-benzyl-
4,6-O-benzylidene-a-D-mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-
b-D-glucopyranoside (18) (91%). Selective cleavage of the
o-nitrobenzyl group on the aglycon moiety of 18 via sunlight
afforded 19 (88%), and subsequent treatment of 19 with the
Dess–Martin reagent afforded the aldehyde 7 (96%).
Results and discussion
In a retrosynthetic study of [125I]2, the phenyl carboxylic acids 3
and 4 were coupled to the trisaccharide derivative 5, which
could be obtained by reductive amination of the primary amine
6 and the aldehyde 7 (Scheme 1). The aldehyde 7, bearing a
trisaccharide composed of N-acetyl-D-glucosamine (GlcNAc), D-
mannose (Man), and D-glucose (Glc), was synthesized via the
thioglycoside glycosylation using odorless benzenethiol as the
activating group.12–15 Successive glycosylation reactions using
p-dodecylphenyl 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-1-thio-
b-D-glucopyranoside (8)13,14, p-dodecylphenyl 2-O-acetyl-3-O-
benzyl-4,6-O-benzylidene-1-thio-a-D-mannopyranoside (9)13,14
and 2-(2-nitrobenzyloxy)ethyl 2,3,4-O-benzyl-b-D-glucopyranoside
,
The amine 6 was synthesized from the commercially
available glycidyl hexadecyl ether in three steps: nucleophilic
attack of the azide ion onto the epoxide of the glycidyl
hexadecyl ether, and subsequent protection of the generated
hydroxyl group with a benzyl group to afford 20, the
{ Uptake amounts of [125I]1a–c to cells were assessed as follows. GnT-V-
transfected Widr cells were seeded at a density of 4 ꢂ 105 per well in 12-
well plates and incubated for 24 h with Roswell Park Memorial Institute hydrogenation of which afforded the desired amine
6
medium (RPMI) 1640 medium containing 10% fetal bovine serum. Afterward,
the medium was replaced by Minimum Essential Medium (MEM) buffer
(0.5 mL/well), and the cells were then pre-incubated for 30 min. After
removing the buffer, MEM buffer solution of [125I]1a–c (18.5 kBq/0.5 mL) was
added, and cells were incubated at 37 ꢃC with 5% CO2. At each time point
(0.5, 1, 3, 6, and 24 h), cells were washed with phosphate-buffered saline
and lysed with 0.2 M NaOHaq. (0.7 mL/well). Radioactivity of the lysate was
measured in Cobra II auto gamma counter (Packard, Detroit, MI, USA). Uptake
profiles were evaluated by the radioactivity, of which the value was corrected
on the basis of the protein concentration measured by bicinchoninic acid
method.
(Scheme 4).
Amine 6 and aldehyde 7 were then coupled by reductive
amination with NaB(CN)H3 to afford the coupling compound
21. The amino group of 21 was protected with a tert-
butyloxycarbonyl (Boc) group to give 22 (54%, two steps).
Removal of the N-phthalimido group with hydrazine hydrate
and subsequent acetylation gave 23. Cleavage of the
benzylidene and benzyl groups of 23, via catalytic
hydrogenolysis followed by acetylation, gave the peracetylated
J. Label Compd. Radiopharm 2013, 56 562–572
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K. Arimitsu et al.
Scheme 1. Retrosynthetic analysis of the 125I-radiolabeled trisaccharide [125I]2.
Scheme 2. Synthesis of the glucose derivative of intermediate 10.
Scheme 3. Synthesis of the trisaccharide intermediate 7.
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Copyright © 2013 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2013, 56 562–572

K. Arimitsu et al.
Scheme 4. Synthesis of compound 2.
Scheme 5. Radiosynthesis of the target compound [125I]2.
trisaccharide derivative 24. The N-tert-butyloxycarbonyl group of compound 2 (82%). The synthesis of the precursor of [125I]2 (26),
24 was cleaved by treatment with methanesulfonic acid in which the tributyltin group was expected to be easily replaced
to afford the secondary amine 5, which was condensed with with a radioiodine atom by iododestannylation, was performed by
15-(4-iodophenyl)pentadecanoic acid (3)19 in the presence of coupling 5 and the carboxylic acid 419 (81%, two steps, Scheme 4).
1-ethyl-3-(30-dimethylaminopropyl)carbodiimideꢀHCl (WSC) and
Next, the tributyltin derivative 26 was transformed to the
DMAP in dichloromethane to give 25 (65%, two steps). 25 was radiolabeled [125I]25 by an iododestannylation reaction using
subsequently deacetylated to produce the nonradioactive hydrogen peroxide as the oxidant.20–22 Conventional
J. Label Compd. Radiopharm 2013, 56 562–572
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K. Arimitsu et al.
deacetylation of [125I]25 afforded the desired radioiodinated and further refined by distillation (180 ꢃC, 2 mmHg) to afford
compound [125I]2. The radiochemical identity of [125I]2 was compound 11 (6.40 g, 70%) as a yellow oil. The NMR spectra of
verified by HPLC with co-injection of the nonradioactive 2. A 11 were in accord with those published.18 1H-NMR (400 MHz,
single radioactive peak of the final radioiodinated compound CDCl3, d): 2.11 (br s, 1H), 3.70–3.72 (m, 2H), 3.82–3.84 (m, 2H),
[
125I]2 was observed at the same retention time as that of the 4.95 (s, 2H), 7.43–7.48 (m, 1H), 7.63–7.67 (m, 1H), 7.77–7.79 (m,
nonradioactive 2. The radioiodinated product was obtained in 1H), 8.06 (dd, J = 1.3, 8.1 Hz, 1H). 13C-NMR (100 MHz, CDCl3) d:
49% radiochemical yield with a radiochemical purity of >95% 61.8, 69.7, 72.3, 124.7, 128.1, 128.7, 133.6, 134.6, 147.4.
after purification by HPLC (Scheme 5).
2-(2-Nitrobenzyloxy)ethyl b-D-glucopyranoside (13)
Conclusions
Boron trifluoride etherate complex (2.57 mL, 20.3mmol) was
We successfully synthesized a novel glycolipid derivative 2 with a
trisaccharyl moiety and a double-chained lipid moiety via the key
intermediate 7, which included a readily modified aglycon
moiety. The 125I-labeled derivative [125I]2 was directly synthesized
from the tributyltin derivative 26 by iododestannylation and
deacetylation. These methods are applicable to the synthesis
and radiosynthesis of various complex glycolipids. In vitro and
in vivo studies using [125I]2 are in progress, and the results will
be published in due course.
added to
a solution of penta-O-acetyl-b-D-glucopyranoside
(7.93 g, 20.3mmol) and 11 (6.01 g, 30.5 mmol) in CH2Cl2 (100 mL)
at 0 ꢃC, and the mixture was stirred for 10 h at room temperature.
The reaction was quenched by pouring the reaction mixture into
ice water, which was then extracted with ethyl acetate. The organic
layer was washed with brine, dried over magnesium sulfate, and
evaporated. The residue was purified using silica gel column
chromatography (n-hexane/ethyl acetate = 2/1) to afford a mixture
of 11 and 2-(2-nitrophenoxy)ethyl 2,3,4,6-tetra-O-acetyl-b-D-
glucopyranoside (12) (10.6g). A solution of the mixture in
methanol (100 mL) was treated with sodium methoxide (28% in
methanol, 4 mL) for 2 h at room temperature. The reaction mixture
was neutralized by adding Dowex 50 (H+) (30.4g), and then the
filtrate was concentrated under vacuum. The residue was purified
using silica gel column chromatography (chloroform/methanol/
Experimental
General
Infrared (IR) spectra were recorded on an FTIR-8300 diffraction grating
infrared spectrophotometer (Shimadzu, Kyoto, Japan). 1H, 13C-NMR
spectra were obtained on a JNM AL-300 spectrometer (JEOL, Tokyo, H2O = 9/1/0.08) to afford compound 13 (5.55 g, 76%, two steps).
1
[a]D = ꢁ14.6ꢃ (c = 1.00, MeOH). H-NMR (400 MHz, C5D5N, d): 3.79
Japan) or unity INOVA-400 spectrometer (Varian, Palo Alto, CA, USA),
and chemical shifts were reported in ppm using tetramethylsilane, CDCl3,
or C5D5N as the internal standard. Mass spectra (MS) were determined on
(br t, J= 5.7 Hz, 2H, –OCH2), 3.94 (ddd, J= 2.4, 5.3, 9.3 Hz, 1H, H-5),
3.98 (br dt, J= 5.7, 11.2Hz, 1H, –OCH2), 4.05 (br t, J= 7.7 Hz, 1H, H-
a JMS-SX 102A QQ or JMS-GC-mate mass spectrometer (JEOL). Specific
2), 4.22–4.28 (m, 2H, H-3, H-4), 4.28 (br dt, J= 5.7, 11.2 Hz, 1H, –
rotations were recorded on a SEPA-200 automatic digital polarimeter
OCH2), 4.38 (dd, J= 5.3, 11.7 Hz, 1H, H-6), 4.55 (dd, J= 2.4, 11.7 Hz,
(Horiba, Kyoto, Japan). Silica gel 60 (70–230 mesh; Merck, Darmstadt,
1H, H-6), 4.91 (d, J= 7.7 Hz, 1H, H-1), 4.95 (s, 2H, –ONB), 6.42 (br,
Germany) was used for open column chromatography. Flash column
chromatography was performed using silica gel 60 (230–400 mesh;
1H, –OH), 7.17–7.32 (m, 3H, –OH), 7.30 (br t, J= 8.1 Hz, 1H), 7.50
(dt, J= 1.3, 7.9 Hz, 1H), 7.90 (dd, J= 1.1, 7.9 Hz, 1H), 8.03 (dd, J= 1.3,
8.1 Hz, 1H). 13C-NMR (100 MHz, C5D5N) d: 62.8 (C-6), 68.9 (–
Merck) as
a solid support for the immobile phase. Thin layer
chromatography (TLC) and preparative TLC were conducted using
Kieselgel 60F-254 plates (0.25 and 0.5 mm; Merck). Unless purification OCH2), 69.8 (–ONB), 70.9 (–OCH2), 71.6 (C-4), 75.2 (C-2), 78.56 (C-
with silica gel gave adequately pure compounds, the compounds were
further purified using a recycling HPLC (LC-908; JAI, Tokyo, Japan)
equipped with a gel permeation chromatography (GPC) column (JAIGEL
1H and 2H, JAI). When possible, diastereomer mixtures were separated
using a recycling HPLC (LC-908) equipped with a silica gel column (Si-
10, 30 mm ꢂ 400 mm; Kusano, Tokyo, Japan), after the purification
methods mentioned earlier.
3), 78.61 (C-5), 104.9 (C-1), 124.0, 124.8, 128.2, 129.2, 133.9, 147.6.
IR (KBr) n: 3393, 2924, 2878, 2361, 2341, 1611, 1522, 1360, 1340,
1298, 1107, 1074 cmꢁ1. FAB-MS m/z: 382 (M + Na+). HRMS m/z:
382.1111 (calcd for C15H21NO9Na: 382.1114). Anal. calcd for
C15H21NO9: C, 50.14; H, 5.89; N, 3.90; found: C, 50.21; H, 5.61; N, 3.83.
2-(2-Nitrobenzyloxy)ethyl 6-O-trityl-b-D-glucopyranoside (14)
Materials
Trityl chloride (8.27 g, 29.7 mmol) and DMAP (catalytic amount)
were added to a solution of 13 (5.33 g, 14.8 mmol) in pyridine
(100 mL), and the mixture was stirred for 6 h under reflux. The
reaction mixture was concentrated in vacuo and poured into
ice water, which was then extracted with chloroform. The
organic layer was washed with brine, dried over magnesium
sulfate, and evaporated. The residue was purified using silica
Most reagents were obtained from Wako Pure Chemical
Industries (Tokyo, Japan), Nacalai Tesque (Kyoto, Japan), and
Aldrich Chemical (St. Louis, MO, USA).
2-(2-Nitrobenzyloxy)ethanol (11)
Ethyleneglycol (100 mL) was added dropwise to sodium hydride gel column chromatography (chloroform/methanol = 60/1) to
(2.42 g, 55.5 mmol) at 0 ꢃC, and the mixture was stirred for 30 min afford 14 (8.03 g, 90%) as an amorphous powder. [a]D = ꢁ27.1ꢃ
1
after elevating the temperature to ca. 20 ꢃC. o-Nitrobenzyl (c = 1.10, CHCl3). H-NMR (400 MHz, CDCl3) d: 3.09 (d, J = 1.8 Hz,
bromide (10.0 g, 46.3 mmol) was added to the reaction mixture, 1H, –OH), 3.36–3.45 (m, 6H), 3.50–3.57 (m, 2H), 3.72–3.84 (m,
which was then stirred for another 5.5 h at room temperature. 3H), 4.05–4.10 (m, 1H), 4.34 (d, J = 7.7 Hz, 1H, H-1), 4.92 (s, 2H),
The reaction was quenched by pouring the mixture into ice 7.22 (tt, J = 1.3, 7.1 Hz, 3H), 7.28 (br t, J = 7.1 Hz, 6H), 7.39 (br t,
water, which was then extracted with ethyl acetate. The organic J = 8.1 Hz, 1H), 7.44 (br d, J = 8.4 Hz, 6H), 7.57 (dt, J = 1.3, 7.9 Hz,
layer was washed with brine, dried over magnesium sulfate, and 1H), 7.75 (dd, J = 1.1, 7.9 Hz, 1H), 8.00 (dd, J = 1.3, 8.1 Hz, 1H).
evaporated. The residue was purified using silica gel column 13C-NMR (100 MHz, CDCl3) d: 64.2 (C-6), 68.8 (–OCH2), 69.8
chromatography (n-hexane/ethyl acetate = 50/1 to ethyl acetate) (–ONB), 70.3 (–OCH2), 71.9 (C-4), 73.5 (C-5), 74.1 (C-2), 76.2
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K. Arimitsu et al.
(C-3), 87.0 (–CPh3), 102.8 (C-1), 124.7, 127.2 (3C), 127.9 (6C), (2C), 128.4 (2C), 128.5 (2C), 128.6, 133.6, 135.0, 137.9, 138.4, 138.5,
128.1, 128.6 (6C), 128.8, 133.6, 134.5, 143.6 (3C), 147.4. IR 147.1. IR (CHCl3) n: 3589, 3032, 3011, 2870, 2401, 1526, 1342,
(CHCl3) n: 3593, 3503, 3028, 3017, 2930, 2883, 1526, 1448, 1234, 1198, 1072cmꢁ1. FAB-MS m/z: 652 (M + Na+). HRMS m/z:
1342, 1067 cmꢁ1. FAB-MS m/z: 624 (M + Na+). HRMS m/z: 652.2523 (calcd for C36H39NO9Na: 652.2517).
624.2206 (calcd for C34H35NO9Na: 624.2210).
2-(2-Nitrobenzyloxy)ethyl 2-O-acetyl-3-O-benzyl-4,6-O-
2-(2-Nitrobenzyloxy)ethyl 2,3,4-O-benzyl-6-O-trityl-b-
D-glucopyranoside (15)
benzylidene-a-D-mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-
b-D-glucopyranoside (16)
A catalytic amount of trimethylsilyl trifluoromethanesulfonate A suspension of 10 (2.03 g, 3.07 mmol), 9 (2.08 g, 3.30 mmol), and
was added to a suspension of 14 (31.0 mg, 51.5 mmol), benzyl molecular sieves 4A (MS 4A) (20 g) in CH2Cl2 (60 mL) was stirred
2,2,2-oxy-trichloroacetimidate (38.5 mL, 0.206 mmol), and for 30 min at ꢁ50 ꢃC. Thereafter, AgOTf (796.6 mg, 3.10 mmol)
molecular sieves 5A (MS 5A) (100 mg) in 1,4-dioxane (2mL). After and N-iodosuccinimide (1.68 g, 7.47 mmol) were added to initiate
being stirred for 1 h at room temperature, the reaction mixture the reaction. After being stirred for another 30 min at ꢁ20 ꢃC, the
was poured into a saturated aqueous sodium bicarbonate solution mixture was filtered through CeliteW. The filtrate was
and extracted with ethyl acetate. The organic layer was washed successively washed with
a saturated aqueous sodium
with brine, dried over magnesium sulfate, and evaporated. The thiosulfate solution and brine, dried over magnesium sulfate,
residue was purified using preparative thin layer silica gel and evaporated. The residue was purified using silica gel column
chromatography (n-hexane/ethyl acetate= 3/1) to afford 15 chromatography (n-hexane/ethyl acetate = 3/1) to afford 16
(41.1 mg, 92%) as an amorphous powder. [a]D = +8.1ꢃ (c = 1.15, (2.62 g, 84%) as an amorphous powder. [a]D = +25.6ꢃ (c = 1.03,
1
1
CHCl3). H-NMR (400 MHz, CDCl3) d: 3.24 (dd, J = 3.8, 10.1 Hz, 1H, CHCl3). H-NMR (400 MHz, CDCl3) d: 2.15 (s, 1H, OAc), 3.39–3.50
H-6), 3.42 (ddd, J = 1.8, 3.8, 9.9 Hz, 1H, H-5), 3.56–3.65 (m, 3H, H-2, (m, 3H, Glc-2, Glc-4, –OCH2), 3.54–3.58 (m, 1H, Man-6), 3.64–
H-3, H-6), 3.81–3.96 (m, 4H, H-4, –OCH2, –OCH2), 4.23–4.28 (m, 1H, 3.83 (m, 6H, Glc-3, –OCH2, Man-5, –OCH2, Glc-6), 3.90 (dt,
–OCH2), 4.37 (d, J = 10.3Hz, 1H, –OCH2), 4.54 (d, J = 7.3Hz, 1H, H- J = 4.6, 10.1 Hz, 1H, Glc-5), 3.96 (dd, J = 3.5, 9.9 Hz, 1H, Man-3),
1), 4.70 (d, J = 10.3 Hz, 1H, –OCH2), 4.78 (d, J= 11.0 Hz, 1H, –OCH2), 4.02–4.07 (m, 2H, Man-4, Man-6), 4.21 (dd, J = 4.4, 9.9 Hz, 1H,
4.79 (d, J=10.6Hz, 1H, –OCH2), 4.90 (d, J= 10.6 Hz, 1H, –OCH2), 4.99 Glc-6), 4.45 (d, J = 7.9 Hz, 1H, Glc-1), 4.50 (d, J = 11.0 Hz, 1H), 4.64
(s, 2H, –ONB), 5.04 (d, J=11.0Hz, 1H, –OCH2), 6.85–6.88 (m, 2H), (d, J = 12.3 Hz, 1H), 4.71 (d, J = 11.4 Hz, 2H), 4.78 (d, J = 10.8 Hz,
7.15–7.39 (m, 23H), 7.43–7.52 (m, 7H), 7.84 (dd, J = 1.1, 7.9Hz, 1H), 1H), 4.83 (s, 3H, Man-1, –OCH2), 4.86 (d, J = 11.0 Hz, 1H), 4.95 (d,
8.05 (dd, J = 1.3, 8.1Hz, 1H). 13C-NMR (100 MHz, CDCl3) d: 62.4 (C- J = 11.0 Hz, 1H), 5.00 (d, J = 11.0 Hz, 1H), 5.43 (dd, J = 1.6, 3.3 Hz,
6), 68.7, 69.8, 70.5, 74.6(C-5), 74.8 (–OBn), 75.0 (–OBn), 75.9 (– 1H, Man-2), 5.61 (s, 1H, PhCH<), 7.19–7.39 (m, 24H), 7.44–7.49
OBn), 77.8 (C-4), 82.5 (C-2), 84.6 (C-3), 86.3 (–CPh3), 103.8 (C-1), (m, 3H), 7.76 (dd, J = 1.1, 7.7 Hz, 1H), 8.04 (dd, J = 1.3, 8.2 Hz, 1H).
124.6, 126.9 (3C), 127.6, 127.6, 127.8 (6C), 128.0 (2C), 128.0 (2C), 13C-NMR (100 MHz, CDCl3) d: 21.0, 63.9 (Man-5), 66.3 (Glc-6),
128.2 (3C), 128.2 (3C), 128.3 (2C), 128.4 (2C), 128.5, 128.8 (5C), 68.6 (Man-6), 68.9 (–OCH2), 69.4 (Man-2), 69.6 (–ONB), 70.4
133.7, 135.2, 137.8, 138.5, 138.6, 143.9 (3C), 147.0. IR (CHCl3) n: (–OCH2), 71.9 (–OBn), 73.4 (Man-3), 74.0 (Glc-5), 74.7 (–OBn),
3088, 3065, 3032, 3011, 2860, 1526, 1448, 1342, 1234, 1094, 75.0 (–OBn), 75.7 (–OBn), 77.7 (Glc-4), 78.2 (Man-4), 82.8
1070 cmꢁ1. FAB-MS m/z: 894 (M+ Na+). HRMS m/z: 894.3622 (calcd (Glc-2), 84.6 (Glc-3), 98.6 (Man-1), 101.4 (PhCH<), 103.6 (Glc-
for C55H53NO9Na: 894.3618).
1), 124.6, 126.0 (2C), 127.6, 127.65 (2C), 127.71 (2C), 127.9
(5C), 128.0 (2C), 128.1 (2C), 128.28 (3C), 128.31 (2C), 128.4
(2C), 128.47 (2C), 128.51, 128.8, 133.6, 135.2, 137.5, 137.8,
137.9, 138.4, 138.5, 147.0, 170.1. IR (CHCl3) n: 3090, 3067,
2-(2-Nitrobenzyloxy)ethyl 2,3,4-O-benzyl-b-
D-glucopyranoside (10)
3030, 3020, 2862, 1746, 1526, 1356, 1234, 1096, 1069 cmꢁ1
.
A solution of 4 M HCl in 1,4-dioxane (4mL) was added to a solution FAB-MS m/z: 1034 (M + Na+). HRMS m/z: 1034.3942 (calcd
of 15 (802.9mg, 0.921mmol) in 1,4-dioxane (28 mL), and the for C58H61NO15Na: 1034.3939).
mixture was stirred for 4 h at room temperature. The reaction
mixture was poured into a saturated aqueous sodium bicarbonate
solution, which was then extracted with ethyl acetate. The organic
layer was washed with brine, dried over magnesium sulfate, and
evaporated. The residue was purified using silica gel column
2-(2-Nitrobenzyloxy)ethyl 3-O-benzyl-4,6-O-benzylidene-a-
D-mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-b-D-
glucopyranoside (17)
chromatography (n-hexane/ethyl acetate= 2/1) to afford 10 Sodium methoxide (28% in methanol, 471 mL) was added to a
(457.1mg, 79%) as an amorphous powder. [a]D = +6.9ꢃ (c = 0.99, solution of 16 (2.39 g, 2.36 mmol) in a mixture of toluene
CHCl3). 1H-NMR (400 MHz, CDCl3) d: 1.96 (br, 1H, –OH), 3.38 (ddd, (10 mL) and methanol (50 mL), and then the mixture was stirred
J = 2.7, 4.6, 9.5Hz, 1H, H-5), 3.46 (dd, J = 7.9, 9.2Hz, 1H, H-2), 3.58 for 1 h at room temperature. The reaction mixture was
(t, J = 9.5Hz, 1H, H-4), 3.66–3.88 (m, 6H, H-3, –OCH2–CH2O–, H-6), neutralized with Dowex 50 (H+) and filtered. The filtrate was
4.08–4.13 (m, 1H, H-6), 4.53 (d, J = 7.9 Hz, 1H, H-1), 4.64 (d, concentrated in vacuo, and the residue was purified using silica
J = 11.0 Hz, 1H), 4.72 (d, J = 11.0Hz, 1H), 4.81 (d, J = 11.0 Hz, 1H), gel column chromatography (n-hexane/ethyl acetate = 2/1) to
4.87 (d, J = 11.0Hz, 1H), 4.93 (d, J = 10.8 Hz, 1H), 4.94 (s, 2H), 4.97 afford 17 (2.08 g, 91%). [a]D = +38.2ꢃ (c = 1.14, CHCl3). 1H-NMR
(d, J = 11.0 Hz, 1H), 7.22–7.35 (m, 15H), 7.38–7.42 (m, 1H), 7.53 (dt, (400 MHz, CDCl3) d: 2.63 (s, 1H, –OH), 3.41–3.49 (m, 3H, Glc-2,
J = 1.3, 7.9 Hz, 1H), 7.79 (dd, J = 1.1, 7.9Hz, 1H), 8.05 (dd, J = 1.3, Glc-4, Glc-5), 3.61–3.69 (m, 2H, Glc-3, –OCH2), 3.73–3.91 (m, 7H,
8.1 Hz, 1H). 13C-NMR (100 MHz, CDCl3) d: 62.0 (C-6), 69.2 (–OCH2), Glc-6, Man-3, Man-5, Man-6, –OCH2), 4.04–4.13 (m, 3H, Man-2,
69.7 (–ONB), 70.3 (–OCH2), 74.8 (–OBn), 75.1 (2C, C-5, –OBn), 75.2 Man-4, –OCH2), 4.22 (dd, J = 3.8, 9.2 Hz, 1H, Man-6), 4.46 (d,
(–OBn), 77.4 (C-4), 82.2 (C-2), 84.4 (C-3), 103.9 (C-1), 124.6, 127.59, J = 7.9 Hz, 1H, Glc-1), 4.54 (d, J = 10.8 Hz, 1H, –OBn), 4.71 (d,
127.62, 127.87 (2C), 127.89, 127.91 (2C), 127.94 (2C), 128.1, 128.3 J = 11.0 Hz, 1H, –OBn), 4.72 (d, J = 12.1 Hz, 1H, –OBn), 4.79
J. Label Compd. Radiopharm 2013, 56 562–572
Copyright © 2013 John Wiley & Sons, Ltd.
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K. Arimitsu et al.
(d, J = 10.8 Hz, 1H, –OBn), 4.84 (d, J = 12.1 Hz, 1H, –OBn), 4.86 3017, 2914, 2870, 1751, 1719, 1526, 1389, 1367, 1238, 1084,
(s, 2H, –ONB), 4.87 (d, J = 10.8 Hz, 1H, –OBn), 4.94 (s, 1H, Man-1), 1040cmꢁ1. FAB-MS m/z: 1409 (M+ Na+). HRMS m/z: 1409.4904
4.95 (d, J = 10.8Hz, 1H, –OBn), 5.00 (d, J = 11.0 Hz, 1H, –OBn), 5.60 (calcd for C76H78N2O23Na: 1409.4893).
(s, 1H, PhCH<), 7.22–7.39 (m, 24H), 7.44–7.50 (m, 3H), 7.77 (dd,
J = 1.1, 7.9Hz, 1H), 8.04 (dd, J = 1.3, 8.1 Hz, 1H). 13C-NMR (100 MHz,
2-Hydroxyethyl 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-b-
CDCl3) d: 63.4 (Man-5), 66.1 (Glc-6), 68.8 (Man-6), 68.9 (–OCH2),
D-glucopyranosyl-(1!2)-3-O-benzyl-4,6-O-benzylidene-a-D-
69.6 (–ONB), 69.8 (Man-2), 70.4 (–OCH2), 72.9 (–OBn), 74.2 (Glc-
mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-b-D-
5), 74.7 (–OBn), 75.1 (–OBn), 75.2 (Man-3), 75.7 (–OBn), 77.6
(Glc-4), 78.8 (Man-4), 82.2 (Glc-2), 84.6 (Glc-3), 100.1 (Man-1),
glucopyranoside (19)
101.5 (PhCH<), 103.7 (Glc-1), 124.6, 126.0 (2C), 127.6, 127.7, A solution of 18 (498.7 mg, 0.359 mmol) in CHCl3 (359 mL) was
127.75, 127.83 (2C), 127.9 (6C), 128.0 (2C), 128.1 (2C), 128.3 irradiated with sunlight for 4 h, and then the solvent was
(2C), 128.4 (2C), 128.45 (2C), 128.49 (2C), 128.6, 128.8, 133.6, evaporated. The residue was purified using silica gel column
135.2, 137.6, 137.86, 137.92, 138.4, 138.5, 147.0. IR (CHCl3) chromatography (n-hexane/ethyl acetate = 2/3) to afford 19
n: 3568, 3090, 3067, 3024, 2916, 2874, 1526, 1454, 1342, (396.6 mg, 88%). [a]D = +15.1ꢃ (c = 1.13, CHCl3). 1H-NMR
1097, 1069, 1030 cmꢁ1. FAB-MS m/z: 992 (M + Na+). HRMS (400 MHz, CDCl3) d: 1.90, 2.05, and 2.06 (each s, 3H, Ac), 2.79 (br
m/z: 992.3837 (calcd for C56H59NO14Na: 992.3833).
t, J = 7.0 Hz, 1H, –OH), 3.04 (t, J = 10.3 Hz, 1H, Man-6), 3.28 (t,
J = 9.7 Hz, 1H, Glc-4), 3.37–3.41 (m, 1H, Glc-5), 3.41 (dd, J = 7.9,
9.2 Hz, 1H, Glc-2), 3.47–3.56 (m, 3H, Man-5, Glc-6), 3.58–3.74 (m,
4H, Glc-3, Man-6, –OCH2), 3.82–3.89 (m, 3H, Man-3, Man-4,
–OCH2), 3.94–4.00 (m, 2H, GlcN-5, –OCH2), 4.19 (br s, 1H,
Man-2), 4.25 (dd, J = 2.6, 12.3 Hz, 1H, GlcN-6), 4.33 (dd,
J = 4.8, 12.3 Hz, 1H, GlcN-6), 4.38 (d, J = 11.0 Hz, 1H, –OBn),
2-(2-Nitrobenzyloxy)ethyl 3,4,6-tri-O-acetyl-2-deoxy-2-
phthalimido-b-D-gluco-pyranosyl-(1!2)-3-O-benzyl-4,6-O-
benzylidene-a-D-mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-
b-D-glucopyranoside (18)
A suspension of 17 (2.46 g, 2.54 mmol), 7 (1.93 g, 2.77 mmol), and 4.42 (d, J = 7.9 Hz, 1H, Glc-1), 4.49 (dd, J = 8.6, 11.0 Hz, 1H,
MS 4A (50 g) in CH2Cl2 (80 mL) was stirred for 30 min at ꢁ50 ꢃC, GlcN-2), 4.58 (d, J = 1.5 Hz, 1H, Man-1), 4.66 (d, J = 12.5 Hz,
and then AgOTf (671.1 mg, 2.61 mmol) and N-iodosuccinimide 1H, –OBn), 4.74 (d, J = 12.5 Hz, 1H, –OBn), 4.77 (d,
(1.43 g, 6.36 mmol) were added to initiate the reaction. After J = 11.0 Hz, 2H, –OBn ꢂ 2), 4.80 (d, J = 11.2 Hz, 1H, –OBn),
being stirred for another 1.5 h at ꢁ20 ꢃC, the mixture was filtered 4.92 (d, J= 11.2 Hz, 1H, –OBn), 4.95 (d, J= 10.8 Hz, 1H, –OBn), 5.20
through CeliteW. The filtrate was neutralized with a saturated (dd, J= 9.2, 10.1Hz, 1H, GlcN-4), 5.43 (s, 1H, PhCH<), 5.46 (d,
aqueous sodium bicarbonate solution, successively washed with J= 8.6 Hz, 1H, GlcN-1), 5.89 (dd, J=9.2, 11.0Hz, 1H, GlcN-3), 7.13–
a saturated aqueous sodium thiosulfate solution and brine, dried 7.41 (m, 25H), 7.69 (br, 2H), 7.84 (br, 2H). 13C-NMR (100 MHz, CDCl3)
over magnesium sulfate, and evaporated. The residue was d: 20.5, 20.7, 20.8, 54.4 (GlcN-2), 62.17 (GlcN-6), 62.23 (–OCH2), 63.9
purified using silica gel column chromatography (n-hexane/ethyl (Man-5), 66.4 (Glc-6), 68.3 (Man-6), 69.2 (GlcN-4), 70.4 (GlcN-3), 71.2
acetate = 2/1) to afford 18 (3.27 g, 93%). [a]D = +19.4ꢃ (c = 1.04, (–OBn), 71.9 (GlcN-5), 73.1 (Man-3), 73.5 (Glc-5), 73.6 (–OCH2), 74.7
1
CHCl3). H-NMR (400 MHz, CDCl3) d: 1.90, 2.02, and 2.06 (each s, (Man-2), 74.9 (–OBn), 75.0 (–OBn), 75.7 (–OBn), 77.5 (Glc-5), 77.9
3H, Ac), 2.97 (t, J = 10.1 Hz, 1H, Man-6), 3.29–3.33 (m, 2H, Glc-4, (Man-4), 82.3 (Glc-2), 84.6 (Glc-3), 96.4 (GlcN-1), 97.8 (Man-1), 101.4
Glc-5), 3.40 (dd, J = 7.9, 9.0 Hz, 1H, Glc-2), 3.49–3.55 (m, 2H, (PhCH<), 104.3 (Glc-1), 123.4, 126.0 (3C), 127.5, 127.7 (3C), 127.76,
Man-5, Man-6), 3.61–3.65 (m, 3H, Glc-3, Glc-6, Man-6), 3.69 127.81 (3C), 127.9 (3C), 128.0 (2C), 128.1 (2C), 128.2 (3C), 128.4 (3C),
(dd, J = 5.1, 8.4 Hz, 1H, –OCH2), 3.78–3.86 (m, 4H, Man-3. Man-4, 128.45 (3C), 128.47 (3C), 128.8, 134.2, 137.5, 137.7, 138.2, 138.28,
–OCH2), 4.00 (ddd, 1H, J = 2.4, 4.8, 10.3 Hz, 1H, GlcN-5), 4.17 (dd, 138.33, 169.5, 170.2, 170.8. IR (CHCl3) n: 3699, 3013, 2864, 2399,
J = 1.6, 2.7Hz, 1H, Man-2), 4.25 (dd, J = 2.4, 12.3 Hz, 1H, GlcN-6), 2363, 2340, 1751, 1719, 1387, 1367, 1238, 1086 cmꢁ1. FAB-MS m/z:
4.27 (dd, J =4.4, 8.4 Hz, 1H, –OCH2), 4.34 (d, J = 11.4Hz, 1H, –OBn), 1274 (M + Na+). HRMS m/z: 1274.4580 (calcd for C69H73NO21Na:
4.35 (dd, J = 4.8, 12.3Hz, 1H, GlcN-6), 4.46 (d, J= 7.9 Hz, 1H, Glc- 1274.4573).
1), 4.50 (dd, J= 8.6, 11.0 Hz, 1H, GlcN-2), 4.62 (d, J= 1.6 Hz, 1H, Man-
1), 4.67 (d, J= 12.5 Hz, 1H, –OBn), 4.70 (d, J= 11.0 Hz, 1H, –OBn),
2-Oxoethyl 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-b-D-
4.73 (d, J=12.5Hz, 1H, –OBn), 4.74 (d, J=11.4Hz, 1H, –OBn), 4.77
glucopyranosyl-(1!2)-3-O-benzyl-4,6-O-benzylidene-a-D-
(d, J= 10.8 Hz, 1H, –OBn), 4.88 (s, 2H, –ONB), 4.94 (d, J= 10.8 Hz, 1H,
mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-b-D-
–OBn), 5.02 (d, J= 11.0 Hz, 1H, –OBn), 5.21 (dd, J=9.2, 10.1Hz, 1H,
glucopyranoside (7)
GlcN-4), 5.41 (s, 1H, PhCH<), 5.45 (d, J = 8.6 Hz, 1H, GlcN-1),
5.90 (dd, J = 9.2, 11.0 Hz, 1H, GlcN-3), 7.10–7.40 (m, 26H), Dess–Martin reagent (51.3 mg, 0.121 mmol) was added to a
7.48 (dt, J = 1.3, 7.9 Hz, 1H), 7.71–7.73 (m, 2H), 7.77 (dd, solution of 19 (100.9 mg, 80.6 mmol) in CH2Cl2 (3 mL), and the
J = 1.1, 7.9 Hz, 1H), 7.84 (br, 2H), 8.04 (dd, J = 1.3, 8.1 Hz, 1H). mixture was stirred for 1 h at room temperature. Subsequently,
13C-NMR (100 MHz, CDCl3) d: 20.5, 20.6, 20.8, 54.4 (GlcN-2), a second amount of Dess–Martin reagent (51.3 mg, 0.121 mmol)
62.2 (GlcN-6), 63.8 (Man-5), 66.5 (Glc-6), 68.3 (Man-6), 69.2 was added to the reaction mixture, which was stirred for another
(GlcN-4), 69.4 (–OCH2), 69.6 (–ONB), 70.3 (GlcN-3), 70.4 (–OCH2), 1 h. The reaction was quenched by pouring it into a saturated
71.2 (–OBn), 72.0 (GlcN-5), 72.7 (Man-3), 74.3 (Glc-5), 74.6 aqueous sodium bicarbonate solution, which was then extracted
(Man-2), 74.7 (–OBn), 74.9 (–OBn), 75.8 (–OBn), 77.1 (Glc-4), with chloroform. The organic layer was successively washed with
78.0 (Man-4), 82.3 (Glc-2), 84.5 (Glc-3), 96.3 (GlcN-1), 98.2 (Man- a saturated aqueous sodium thiosulfate solution and brine, dried
1), 101.3 (PhCH<), 104.0 (Glc-1), 123.4, 124.6, 126.0 (2C), over sodium sulfate, and evaporated. The residue was purified
127.58, 127.61, 127.7 (3C), 127.79 (4C), 127.83, 127.91 (3C), 127.94 using silica gel column chromatography (n-hexane/ethyl
(3C), 128.0 (3C), 128.26 (2C), 128.29 (2C), 128.36 (2C), 128.43 (2C), acetate = 1/1) to afford 7 (96.8 mg, 96%). [a]D = +21.7ꢃ (c = 0.98,
1
128.5 (3C), 128.7, 133.6, 134.2, 135.1, 137.6, 137.8, 138.2, 138.3, CHCl3). H-NMR (400 MHz, CDCl3) d: 1.90 (s, 3H, Ac), 2.06 (s, 6H,
138.4, 147.0, 169.4, 170.2, 170.7. IR (CHCl3) n: 3481, 3067, 3030, Ac x2), 3.03 (t, J = 10.3 Hz, 1H, Man-6), 3.31–3.33 (m, 2H, Glc-4,
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J. Label Compd. Radiopharm 2013, 56 562–572

K. Arimitsu et al.
Glc-5), 3.45–3.51 (m, 2H, Man-5, Glc-6), 3.47 (dd, J = 7.9, 9.2 Hz, 2-(Benzyloxy)-3-(hexadecyloxy)propanamine (6)
1H, Glc-2), 3.57–3.67 (m, 3H, Glc-3, Glc-6, Man-6), 3.78 (dd,
A catalytic amount of Pd–C was added to a solution of 20
J = 2.9, 10.1 Hz, 1H, Man-3), 3.85 (t, J = 10.1 Hz, 1H, Man-4), 4.01
(20.4 mg, 47.3 mmol) in methanol (1 mL), and the mixture was
(ddd, J = 2.4, 4.8, 10.1 Hz, 1H, GlcN-5), 4.13 (dd, J = 1.5, 2.9 Hz,
stirred for 1 h under hydrogen atmosphere. The mixture was
1H, Man-2), 4.27 (dd, J = 2.4, 12.2 Hz, 1H, GlcN-6), 4.30 (br d,
filtered through CeliteW, and the filtrate was concentrated under
J = 17.8 Hz, 1H, –OCH2), 4.34 (d, J = 11.0 Hz, 1H, –OBn), 4.36 (dd,
vacuum. The residue was purified using silica gel column
J = 4.8, 12.2 Hz, 1H, GlcN-6), 4.45 (d, J = 7.9 Hz, 1H, Glc-1), 4.49
chromatography (chloroform/methanol = 10/1) to afford
6
(dd, J = 8.4, 11.0 Hz, 1H, GlcN-2), 4.56 (d, J = 1.5 Hz, 1H, Man-1),
4.57 (dd, J = 1.3, 17.8 Hz, 1H, –OCH2), 4.69 (d, J= 12.5 Hz, 1H, –
OBn), 4.74 (d, J = 11.0 Hz, 1H, –OBn), 4.75 (d, J = 12.5 Hz, 1H, –OBn),
4.79 (d, J = 11.0 Hz, 1H, –OBn), 4.80 (d, J = 10.8 Hz, 1H, –OBn),
4.97 (d, J = 10.8 Hz, 1H, –OBn), 5.03 (d, J = 11.0 Hz, 1H, –OBn),
5.20 (dd, J = 9.2, 10.1 Hz, 1H, GlcN-4), 5.42 (d, J = 8.4 Hz, 1H,
GlcN-1), 5.43 (s, 1H, PhCH<), 5.85 (dd, J = 9.2, 11.0 Hz, 1H,
GlcN-3), 7.12–7.42 (m, 25H), 7.70 (br, 2H), 7.84 (br, 2H), 9.68
(br d, J = 0.7 Hz, 1H, –CHO). 13C-NMR (100 MHz, CDCl3) d:
20.5, 20.7, 20.8, 54.4 (GlcN-2), 62.2 (GlcN-6), 63.9 (Man-5),
65.9 (Glc-6), 68.3 (Man-6), 69.1 (GlcN-4), 70.4 (GlcN-3), 71.3
(–OBn), 71.9 (GlcN-5), 72.8 (Man-3), 73.8 (Glc-5), 74.85 (Man-
2), 74.92 (2C, –OBn, –OCH2), 75.1 (–OBn), 75.8 (–OBn), 77.1
(Glc-4), 78.0 (Man-4), 82.0 (Glc-2), 84.3 (Glc-3), 96.4 (GlcN-1),
97.8 (Man-1), 101.4 (PhCH<), 103.9 (Glc-1), 123.4, 126.0 (2C),
127.6, 127.7 (2C), 127.77 (3C), 127.84, 127.91, 127.94 (2C),
128.1 (3C), 128.15 (3C), 128.24 (3C), 128.42 (3C), 128.44 (4C),
128.8, 131.5, 134.2 (2C), 137.5, 137.7, 138.1, 138.2, 138.3,
169.5, 170.2, 170.8, 199.3. IR (CHCl3) n: 3067, 3028, 3018,
2858, 1749, 1719, 1603, 1387, 1367, 1236, 1088 cmꢁ1. FAB-
MS m/z: 1272 (M + Na+). HRMS m/z: 1272.4420 (calcd for
(18.9 mg, 98%). 1H-NMR (300 MHz, CDCl3) d: 0.88 (t, J = 6.9 Hz,
3H), 1.25 (m, 28H), 1.52–1.59 (m, 2H), 2.78 (dd, J = 5.9, 13.3 Hz,
1H), 2.88 (dd, J = 3.6, 13.3 Hz), 3.41–3.56 (m, 5H), 4.59
(d, J = 11.7 Hz, 1H), 4.73 (d, J = 11.7 Hz, 1H), 7.26–7.36 (m, 5H).
13C-NMR (75 MHz, CDCl3) d: 14.1, 22.7, 26.1, 29.4, 29.5, 29.61,
29.62, 29.65, 29.68 (3C), 29.69 (3C), 31.9, 43.5, 71.4, 71.7, 72.1,
79.6, 127.6, 127.8 (2C), 128.4 (2C), 138.7. IR (CHCl3) n: 3385,
3003, 2928, 2855, 1585, 1454, 1352, 1113, 1070 cmꢁ1. EI-MS
(20 eV) m/z: 405 (M+). HRMS m/z: 405.3612 (calcd for C26H47NO2:
405.3607).
2-[N-tert-Butoxycarbonyl-N-(2-benzyloxy-3-hexadecyloxy)
propyl]aminoethyl 3,4,6-tri-O-acetyl-2-deoxy-2-phthalimido-
b-D-glucopyranosyl-(1!2)-3-O-benzyl-4,6-O-benzylidene-a-
D-mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-b-D-
glucopyranoside (22)
Sodium cyanoborohydride (88.6 mg, 1.41 mmol) was added to a
solution of 7 (1.70 g, 1.36 mmol) and 6 (672.3 mg, 1.66 mmol) in
CH2Cl2 (70 mL), and the mixture was stirred for 4 h at room
temperature. The reaction was quenched by pouring the mixture
into a saturated aqueous sodium bicarbonate solution, which
was then extracted with chloroform. The organic layer was
washed with brine, dried over sodium sulfate, and evaporated.
Boc2O (1.56 ml, 6.79 mmol) and DMAP (a catalytic amount) were
added to a solution of the residue in CH2Cl2 (50 mL), and the
reaction mixture was stirred for 7 h at room temperature. The
reaction was quenched by pouring the mixture into a saturated
aqueous sodium bicarbonate solution, which was then extracted
with ethyl acetate. The organic layer was washed with brine,
dried over magnesium sulfate, and evaporated. The residue was
purified using silica gel column chromatography (n-hexane/ethyl
acetate = 2/1) to afford 22 (1.29 g, 54%). The two diastereomers of
22 were separated by HPLC (n-hexane/ethyl acetate = 3/2).
C
69H71NO21Na: 1272.4416).
1-Azide-2-benzyloxypropyl hexadecyl ether (20)
Sodium azide (69.8 mg, 1.07 mmol) and triethylamine
hydrochloride (147.7 mg, 1.07 mmol) were added to a solution
of glycidyl hexadecyl ether (160.2 mg, 0.537 mmol) in DMF
(8 mL), and the mixture was stirred for 5 h at 100 ꢃC. The reaction
mixture was poured into
a saturated aqueous sodium
bicarbonate solution, which was then extracted with ethyl
acetate. The organic layer was washed with distilled water, dried
over magnesium sulfate, and evaporated. The residue was
purified using silica gel column chromatography (n-hexane/ethyl
acetate = 7/1) to afford a crude mixture containing 1-azido-3-
(hexadecyloxy)-2-propanol
(31.3 mg, 0.717 mmol) and benzyl bromide (80 mL, 0.673 mmol)
(156.4 mg).
Sodium
hydride
22 (diastereomer A)
were added to a solution of the mixture (151.9 mg) in DMF [a]D = +4.2ꢃ (c = 1.20, CHCl3). 1H-NMR (400 MHz, C5D5N) d: 0.86 (t,
(5 mL) at 0 ꢃC. The mixture was stirred for 2 h at room J = 7.0 Hz, 3H), 1.24–1.64 (m, 37H), 1.86, 2.02, and 2.04 (each s, 3H,
temperature, and the reaction was quenched by pouring the Ac), 3.22 (br t, J = 10.1 H, 1H, Man-6), 3.45–3.49 (m, 2H), 3.60–4.03
mixture into ice water, which was then extracted with diethyl (m, 15H), 4.15–4.38 (m, 4H), 4.46 (dd, J = 2.0, 12.3 Hz, 1H, GlcN-6),
ether. The organic layer was washed with distilled water, dried 4.63–4.78 (m, 5H), 4.85–5.04 (m, 7H), 5.11 (m, 2H), 5.18–5.24 (m,
over magnesium sulfate, and evaporated. The residue was 2H), 5.52 (s, 1H, PhCH<), 5.63 (dd, J = 9.3, 10.1 Hz, 1H, GlcN-4),
purified using silica gel column chromatography (n-hexane/ethyl 6.03 (m, 1H, GlcN-1), 6.47 (dd, J = 9.3, 10.8 Hz, 1H, GlcN-3), 7.17–
1
acetate = 50/1) to afford 20 (145.9 mg, 76%, two steps). H-NMR 7.22 (m, 1H), 7.27–7.44 (m, 22H), 7.51–7.59 (m, 9H), 7.92 (br,
(300 MHz, CDCl3) d: 0.88 (t, J = 6.9 Hz, 3H), 1.25 (m, 26H), 1.53 2H). 13C-NMR (100 MHz, C5D5N) d: 14.3, 20.3, 20.5, 20.7, 22.9,
(m, 2H), 3.38 (dd, J = 6.0, 12.9 Hz, 1H), 3.41 (dd, J = 4.3, 12.9 Hz, 26.6, 28.4, 28.6, 29.6, 29.85, 29.93, 30.0, 30.2, 32.1, 48.4, 48.8,
1H), 3.42 (t, J = 6.6 Hz, 2H), 3.49 (dd, J = 6.0, 10.0 Hz, 1H), 3.54 50.0, 55.4 (GlcN-2), 62.5 (GlcN-6), 64.5 (Man-5), 67.3 (Glc-6), 68.1,
(dd, J = 5.0, 10.0 Hz, 1H), 3.72 (br quint, J = 6.0 Hz, 1H), 4.68 (s, 68.7 (Man-6), 69.8 (GlcN-4), 70.9, 71.0 (GlcN-3), 71.8 (GlcN-5),
2H), 7.27–7.37 (m, 5H). 13C-NMR (75 MHz, CDCl3) d: 14.1, 22.7, 72.4 (Glc-4), 72.5 (Man-3), 74.1, 74.4, 74.8 (Man-2), 75.7, 77.8
26.1, 29.4, 29.5, 29.60 (2C), 29.62, 29.65, 29.66, 29.68, 29.69 (3C), (Glc-5), 78.0, 78.8 (Man-4), 79.4, 82.7 (Glc-2), 85.1 (Glc-3), 96.7
31.9, 52.1, 70.2, 71.8, 72.3, 77.1, 127.78, 127.82 (2C), 128.4 (2C), (GlcN-1), 98.7 (Man-1), 102.1 (PhCH<), 104.2 (Glc-1), 123.99,
138.0. IR (CHCl3) n: 3007, 2926, 2855, 2104, 1711, 1364 cmꢁ1
.
124.04, 126.9, 127.87, 127.91, 127.98, 128.03, 128.1, 128.2, 128.4,
FAB-MS m/z: 454 (M+Na+). HRMS m/z: 454.3415 (calcd for 128.5, 128.6, 128.69, 128.73, 128.8, 129.1, 132.2, 134.7, 136.0,
C26H45N3O2Na: 454.3409).
138.5, 139.0, 139.1, 139.4, 139.6, 139.9, 150.3, 155.7, 168.1,
J. Label Compd. Radiopharm 2013, 56 562–572
Copyright © 2013 John Wiley & Sons, Ltd.
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K. Arimitsu et al.
}
169.2, 169.8, 170.4, 170.5. IR (CHCl3) n: 3015, 2928, 2855, 1749, m/z: 1673 (M + Na+). HRMS m/z: 1673.8655 (calcd for
1719, 1688, 1387, 1367, 1226, 1200, 1088 cmꢁ1. FAB-MS m/z: C94H126N2O23Na: 1673.8649). Anal. calcd for C94H126N2O23: C,
1761 (M + Na+). HRMS m/z: 1761.8595 (calcd for 68.34; H, 7.69; N, 1.70; found: C, 68.12; H, 7.70; N, 1.43.
C100H126N2O24Na: 1761.8598).
2-[N-tert-Butoxycarbonyl-N-(2-acetoxy-3-hexadecyloxy)
22 (diastereomer B)
propyl]aminoethyl 3,4,6-tri-O-acetyl-2-acetamido-deoxy-2-
1
[a]D = +18.8ꢃ (c = 0.95, CHCl3). H-NMR (400 MHz, C5D5N) d: 0.85
b-D-glucopyranosyl-(1!2)-3,4,6-O-acetyl-a-D-
(t, J = 7.0 Hz, 3H), 1.24–1.61 (m, 37H), 1.85, 2.02, and 2.04 (each
mannopyranosyl-(1!6)-2,3,4-tri-O-acetyl-b-D-
s, 3H, Ac), 3.23 (m, 1H, Man-6), 3.45–3.47 (m, 2H), 3.60–4.02 (m,
15H), 4.14–4.39 (m, 4H), 4.46 (dd, J = 2.0, 12.3 Hz, 1H, GlcN-6),
4.63–4.79 (m, 5H), 4.86–5.05 (m, 7H), 5.12 (br d, J = 10.6 Hz, 2H),
5.18–5.26 (m, 2H), 5.52 (s, 1H, PhCH<), 5.63 (t, J = 9.7 Hz, 1H,
GlcN-4), 6.03 (br t, J = 8.4 Hz, 1H, GlcN-1), 6.47 (t, J = 10.1 Hz, 1H,
GlcN-3), 7.20–7.22 (m, 1H), 7.27–7.44 (m, 22H), 7.52–7.60 (m,
9H), 7.93 (br, 2H). 13C-NMR (100 MHz, C5D5N) d: 14.3, 20.3, 20.5,
20.7, 22.9, 26.6, 28.4, 28.6, 29.6, 29.8, 29.9, 29.98, 30.01, 30.18,
32.1, 48.7, 48.8, 50.5, 55.4 (GlcN-2), 62.5 (GlcN-6), 64.5 (Man-5),
67.3 (Glc-6), 68.3, 68.7 (Man-6), 69.8 (GlcN-4), 70.9, 71.0 (GlcN-3),
71.8 (GlcN-5), 72.0, 72.4 (Glc-4), 72.5 (Man-3), 74.0, 74.3, 74.6,
74.8 (Man-2), 75.7, 75.9, 77.9 (Glc-5), 78.0, 78.8 (Man-4), 79.4,
82.6 (Glc-2), 85.1 (Glc-3), 96.7 (GlcN-1), 98.7 (Man-1), 102.1
(PhCH<), 104.1 (Glc-1), 124.0, 124.1, 126.9, 127.9, 127.98,
128.04, 128.1, 128.2, 128.4, 128.6, 128.69, 128.73, 128.8, 129.1,
132.2, 134.7, 136.0, 138.5, 138.8, 139.0, 139.1, 139.4, 139.7,
139.8, 150.3, 155.6, 169.9, 170.4, 170.5.} IR (CHCl3) n: 3026, 3017,
2928, 2855, 1751, 1719, 1686, 1387, 1367, 1236, 1200,
1088 cmꢁ1. FAB-MS m/z: 1761 (M + Na+). HRMS m/z: 1761.8593
(calcd for C100H126N2O24Na: 1761.8598).
glucopyranoside (24)
Pd(OH)2–C (291.6 mg) was added to a solution of 23 (561.9 mg,
0.340 mmol) in methanol (15 mL), and the mixture was stirred
for 15 h under hydrogen atmosphere. The reaction mixture was
filtered through CeliteW, and the filtrate was concentrated
under vacuum. Acetic anhydride (321 mL, 3.40 mmol) and
DMAP (a catalytic amount) were added to a solution of the
residue in pyridine (15 mL), and the mixture was stirred for
4 h. Subsequently, a second quantity of acetic anhydride
(321 mL, 3.40 mmol) was added to the reaction mixture, which
was then stirred for another 5 h. The reaction was quenched
by pouring the mixture into ice water, which was then
extracted with ethyl acetate. The organic layer was washed
with brine, dried over magnesium sulfate, and evaporated.
The residue was purified using silica gel column
chromatography (n-hexane/ethyl acetate = 1/4) to afford 24
(459.2 mg, 96%, two steps). 1H-NMR (400 MHz, C5D5N) d:
0.85 (t, J = 7.1 Hz, 3H), 1.27–1.35 (m, 38H), 1.51–1.59 (m, 2H),
1.97 (s, 6H), 1.99, 2.00, 2.01, 2.06, 2.07, and 2.09 (each s,
3H), 2.10 (br, 9H), 3.46 (br, 2H), 3.66–4.07 (m, 8H), 4.24–4.27
(m, 2H), 4.34 (br, 1H), 4.48 (br, 2H), 4.58–4.62 (m, 1H), 4.75
(br, 1H), 4.90–4.99 (m, 2H), 5.24 (m, 1H), 5.38–5.74 (m, 7H),
5.87 (m, 1H), 6.03 (br, 1H), 9.20 (br, 1H). FAB-MS m/z: 1429
(M + Na+). HRMS m/z: 1429.6724 (calcd for C66H106N2O30Na:
1429.6728).
2-[N-tert-Butoxycarbonyl-N-(2-benzyloxy-3-hexadecyloxy)
propyl]aminoethyl 3,4,6-tri-O-acetyl-2-acetamido-deoxy-2-
b-D-glucopyranosyl-(1!2)-3-O-benzyl-4,6-O-benzylidene-a-
D-mannopyranosyl-(1!6)-2,3,4-tri-O-benzyl-b-D-
glucopyranoside (23)
Hydrazine hydrate (80% in water, 1.0 mL) was added to a
solution of 22 (822.2 mg, 0.473 mmol) in ethanol (15 mL), and
the mixture was stirred for 1.5 h at 80 ꢃC. After cooling the
reaction mixture to room temperature, the precipitate was
filtered, and the filtrate was evaporated. Acetic anhydride
(223 mL, 2.36 mmol) and DMAP (a catalytic amount) were added
to a solution of the filtrate residue in pyridine (15 mL). The
reaction mixture was stirred for 10 h poured into ice water and
extracted with ethyl acetate. The organic layer was washed with
brine, dried over magnesium sulfate, and evaporated. The
residue was purified using silica gel column chromatography
(n-hexane/ethyl acetate = 1/1) to afford 23 (680.5 mg, 87%, 2
2-[N-(2-Acetoxy-3-hexadecyloxy)propyl-15-(4-iodophenyl)
pentadecanecarboxamido]ethyl 3,4,6-tri-O-acetyl-2-
acetamido-deoxy-2-b-D-glucopyranosyl-(1!2)-3,4,6-O-
acetyl-a-D-mannopyranosyl-(1!6)-2,3,4-tri-O-acetyl-b-D-
glucopyranoside (25)
Methanesulfonic acid (one drop) was added to a solution of
24 (177.6 mg, 0.126 mmol) in CH2Cl2 (5 mL), and the mixture
was stirred for 1 h at room temperature. Subsequently, a
further two drops of methanesulfonic acid were added to
the mixture, which was then stirred for another 2.5 h. The
reaction mixture was poured into
a saturated aqueous
1
steps). H-NMR (400 MHz, C5D5N) d: 0.85 (t, J = 7.0 Hz, 3H), 1.23–
sodium bicarbonate solution, which was then extracted with
chloroform. The organic layer was washed with brine, dried
over sodium sulfate, and evaporated.
1.64 (m, 40H), 1.96 (s, 3H), 2.01 (s, 6H), 2.17 (s, 3H), 3.45–5.23
(m, 34H), 5.44–5.54 (m, 3H), 5.59 (s, 1H), 6.00 (br, 1H), 7.19–7.63
(m, 30H), 9.03–9.12 (m, 1H). 13C-NMR (100 MHz, C5D5N) d: 14.3,
20.55, 20.58, 20.64, 22.9, 23.4, 26.6, 28.4, 28.5, 29.6, 29.85, 29.92,
29.98, 30.00, 30.01, 30.2, 32.1, 48.7, 50.1, 55.6, 62.7, 65.1, 67.2,
68.1, 69.1, 69.8, 70.9, 71.8, 72.1, 72.3, 72.9, 74.5, 74.8, 75.0, 75.2,
75.7, 78.0, 78.7, 79.4, 82.7, 85.1, 99.5, 100.4, 102.1, 104.1, 123.0,
124.0, 126.9, 127.8, 127.9, 128.0, 128.1, 128.2, 128.35, 128.43,
128.6, 128.70, 128.73, 128.8, 129.1, 138.7, 139.1, 139.3, 139.5,
155.7, 169.9, 170.5, 170.6, 170.9.} IR (CHCl3) n: 3352, 3011, 2928,
2855, 1751, 1674, 1454, 1367, 1236, 1070, 1045 cmꢁ1. FAB-MS
1-Ethyl-3-(30-dimethylaminopropyl)carbodiimideꢀHCl (WSC;
38.8 mg, 0.202 mmol) and DMAP (a catalytic amount) were
added to a solution of the residue (162.2 mg) and 3 (68.0 mg,
0.153 mmol) in CH2Cl2 (5 mL), and the mixture was stirred for
5 h. The reaction was quenched by pouring the reaction
mixture into
a saturated aqueous ammonium chloride
solution, which was then extracted with ethyl acetate. The
organic layer was washed with brine, dried over sodium
sulfate, and evaporated. The residue was purified using silica
gel column chromatography (chloroform/methanol = 40/1)
and further refined by GPC to afford 25 (141.7 mg, 65%, 2
steps). 1H-NMR (400 MHz, C5D5N) d: 0.86 (t, J = 6.8 Hz, 3H),
} The signals could not be assigned due to the presence of conformers at the
appropriate temperature for measuring the 13C-NMR spectra.
www.jlcr.org
Copyright © 2013 John Wiley & Sons, Ltd.
J. Label Compd. Radiopharm 2013, 56 562–572

K. Arimitsu et al.
1.26–1.66 (m, 55H), 1.85 (br, 3H), 1.96 and 1.97 (each s, J = 10.3 Hz, 0.5H), 6.01 and 6.03 (each t, J = 9.3 Hz, 0.5H), 7.37 (d,
1.5H), 2.00 (br, 6H), 2.07 and 2.08 (each s, 1.5H), 2.10 and J = 7.5 Hz, 2H), 7.63 (d, J = 7.5 Hz, 2H), 9.18 (d, J = 8.4 Hz, 0.5H),
2.12 (each br, 6H), 2.13 and 2.16 (each br, 3H), 2.45–2.68 9.19 (d, J = 8.2 Hz, 0.5H). FAB-MS m/z: 1919 (M + Na+). HRMS m/
(m, 4H), 3.49 (m, 3H), 3.70–4.14 (m, 10H), 4.25–4.37 (m, z: 1919.9723 (calcd for C94H156N2O29SnNa: 1919.9713).
4H), 4.49 (m, 3H), 4.61 (dd, J = 5.5, 12.3 Hz, 1H), 4.76 and
4.79 (each br, 0.5H), 4.95 (d, J = 8.1 Hz, 1H), 5.25 and 5.29
2-[N-(2-Hydroxy-3-hexadecyloxy)propyl-15-(4-iodophenyl)
pentadecanecarboxamido]ethyl 2-acetamido-2-deoxy-b-D-
glucopyranosyl-(1!2)-a-D-mannopyranosyl-(1!6)-b-D-
glucopyranoside (2)
(each s, 0.5H), 5.38–5.63 (m, 5H), 5.70–5.76 (m, 1H), 5.87
and 5.89 (each t, J = 10.1 Hz, 0.5H), 6.01 (t, J = 10.1 Hz,
0.5H), 6.03–6.08 (m, 0.5H), 6.98 (d, J = 7.9 Hz, 2H), 7.71 (d,
J = 7.9 Hz, 2H), 9.17–9.25 (m, 1H). FAB-MS m/z: 1755
(M + Na+). HRMS m/z: 1755.7621 (calcd for C82H129IN2O29Na:
1755.7623).
Sodium methoxide (28% in methanol, 49 mL, 246 mmol) was added
to a solution of 25 (141.1 mg, 81.6 mmol) in methanol (5 mL), and
the mixture was stirred for 30 min at room temperature. The
reaction mixture was neutralized with Dowex 50 (H+) and filtered.
The filtrate was concentrated under vacuum, and the residue was
purified using silica gel column chromatography (chloroform/
methanol/water = 8/2/0.1) to afford 2 (88.1 mg, 82%). The two
diastereomers of 2 were separated by HPLC (using methanol).
2-[N-(2-Acetoxy-3-hexadecyloxy)propyl-15-(4-
tributylstannylphenyl)pentadecanecarboxamido]ethyl 3,4,6-
tri-O-acetyl-2-acetamido-deoxy-2-b-D-glucopyranosyl-
(1!2)-3,4,6-O-acetyl-a-D-mannopyranosyl-(1!6)-2,3,4-tri-
O-acetyl-b-D-glucopyranoside (26)
Methanesulfonic acid (three drops) was added to a solution of 24
(166.6 mg, 0.118 mmol) in CH2Cl2 (5 mL), and the mixture was
stirred for 3.5 h at room temperature. The reaction mixture was
poured into a saturated aqueous sodium bicarbonate solution,
which was then extracted with chloroform. The organic layer
was washed with brine, dried over sodium sulfate, and
evaporated. WSC (37.0 mg, 0.193 mmol) and DMAP (a catalytic
amount) were added to a solution of the residue (152.3 mg)
and 4 (86.7 mg, 0.143 mmol) in CH2Cl2 (5 mL), and the mixture
was stirred for 5 h at room temperature. The reaction mixture
was poured into a saturated aqueous ammonium chloride
solution, which was then extracted with ethyl acetate. The
organic layer was washed with brine, dried over magnesium
sulfate, and evaporated. The residue was purified using silica
gel column chromatography (chloroform/methanol = 40/1) and
further refined by GPC to afford 26 (181.5 mg, 81%, two steps).
The two diastereomers of 26 were separated by HPLC (using
ethyl acetate).
2 (diastereomer A)
1H-NMR (400 MHz, C5D5N) d: 0.85 (t, J = 7.0 Hz, 3H), 1.24–1.33 (m,
46H), 1.52–1.65 (m, 4H), 1.75–1.85 (m, 2H), 2.19 (s, 1.2H), 2.20 (s,
1.8H), 2.47 (br t, J = 6.4 Hz, 2H), 2.54–2.64 (m, 1.2H), 2.71–2.76
(m, 0.8H), 3.46–3.49 (m, 3H), 3.59–3.70 (m, 3H), 3.76–4.64 (m,
22H), 4.86 (d, J = 7.9 Hz, 1H), 5.22 (m, 1H), 5.28 (br d, J = 7.5 Hz,
1H), 5.54 (br s, 1H), 5.64 (br,, 10H), 6.98 (d, J = 8.2 Hz, 2H), 7.71
(d, J = 8.2 Hz, 2H), 8.94 (br, 1H). FAB-MS m/z: 1335 (M + Na+).
HRMS m/z: 1335.6562 (calcd for C62H109IN2O19Na: 1335.6567).
2 (diastereomer B)
1H-NMR (400 MHz, C5D5N) d: 0.85 (t, J = 7.1Hz, 3H), 1.23–1.33 (m,
46H), 1.52–1.63 (m, 4H), 1.75–1.85 (m, 2H), 2.19 (s, 1.2H), 2.20 (s,
1.8H), 2.47 (br t, J = 7.1Hz, 2H), 2.57 (quint, J = 7.7Hz, 1.2H), 2.75
(quint, J = 7.7Hz, 0.8H), 3.46–3.51 (m, 3H), 3.59–4.64 (m, 25H), 4.79
(d, J = 7.9 Hz, 0.6H), 4.83 (d, J = 7.9 Hz, 0.4H), 5.21 (br, 12H), 5.54 (d,
J = 1.5 Hz, 0.6H), 5.55 (d, J = 1.5 Hz, 0.4H), 6.98 (d, J = 8.2Hz, 2H),
7.71 (d, J = 8.2 Hz, 2H), 8.91 (m, 1H). FAB-MS m/z: 1335 (M + Na+).
HRMS m/z: 1335.6561 (calcd for C62H109IN2O19Na: 1335.6567).
26 (diastereomer A)
1H-NMR (400 MHz, C5D5N) d: 0.87 (t, J = 7.3 Hz, 12H), 1.04–1.19 (m,
6H), 1.26–1.41 (m, 52H), 1.52–1.65 (m, 12H), 1.84 (br, 3H), 1.96
and 1.97 (each s, 1.5H), 1.99 (s, 1.5H), 2.01 (br, 4.5H), 2.07 (br,
3H), 2.10–2.13 (m, 12H), 2.16 (m, 6H), 2.56 (br t, J = 7.5 Hz, 1H),
2.62 (m, 3H), 3.50 (m, 3H), 3.71–4.14 (m, 10H), 4.25–4.37 (m,
4H), 4.50 (m, 3H), 4.61 (dd, J = 5.5, 12.6 Hz, 1H), 4.76 and 4.79
(each br, 0.5H), 4.99 (br, 1H), 5.25 and 5.30 (each s, 0.5H), 5.41–
5.50 (m, 4H), 5.58–5.63 (m, 1H), 5.70–5.76 (m, 1H), 5.85–5.92 (m,
1H), 5.99–6.08 (m, 1H), 7.38 (d, J = 7.7 Hz, 2H), 7.64 (d, J = 7.7 Hz,
2H), 9.19 (br d, J = 7.7 Hz, 0.5H), 9.25 (br d, J = 7.9 Hz, 0.5H). FAB-
MS m/z: 1919 (M + Na+). HRMS m/z: 1919.9722 (calcd for
C94H156N2O29SnNa: 1919.9713).
2-[N-(2-Hydroxy-3-hexadecyloxy)propyl-15-(4-[125I]
iodophenyl)pentadecanecarboxamido]ethyl 2-acetamido-2-
deoxy-b-D-glucopyranosyl-(1!2)-a-D-mannopyranosyl-
(1!6)-b-D-glucopyranoside ([125I]2)
Hydrogen peroxide (5% in water, 10 mL), 0.1M hydrochloric acid
(10 mL), and [125I]NaI (5 mL, 13.4MBq) were added to a solution of
tributyltin derivative 26 (100 mg) in acetonitrile (20mL). After stirring
the reaction mixture at room temperature for 1 h, the mixture was
added to a saturated aqueous sodium thiosulfate solution, which
was then extracted with ethyl acetate and concentrated under
26 (diastereomer B)
1H-NMR (400 MHz, C5D5N) d: 0.88 (t, J = 7.3 Hz, 12H), 1.04–1.21 (m, vacuum. The residue was treated with methanol (20mL) and
6H), 1.25–1.43 (m, 52H), 1.54–1.70 (m, 12H), 1.85 (br, 2H), 1.96, sodium methoxide (28% in methanol, 3 mL) for 30 min, and then
1.99, and 1.97 (each s, 1.5H), 2.00 (s, 3H), 2.01, 2.06, and 2.07 the mixture was neutralized with 1 M hydrochloric acid. The
(each s, 1.5H), 2.09–2.12 (m, 12H), 2.125, 2.132, 2.15, and 2.16 residue was purified using HPLC on a Cosmosil 5C18 AR-300
(each s, 1.5H), 2.54–2.68 (m, 4H), 3.44–3.54 (m, 3H), 3.68–4.15 column (4.6mmꢂ 150mm, Nacalai Tesque, Kyoto, Japan) using
(m, 10H), 4.24–4.37 (m, 4H), 4.46–4.55 (m, 3H), 4.61 (dd, J = 5.5, methanol/water (98/2, v/v) as the mobile phase at a flow rate of
12.3 Hz, 1H), 4.76 and 4.79 (each dd, J = 1.6, 3.5 Hz, 0.5H), 4.93 1.0mL/min. [125I]2 (6.6MBq) was isolated, and it was identified by
(d, J = 8.1 Hz, 1H), 5.24 and 5.28 (each d, J = 1.6 Hz, 0.5H), 5.38– co-injection HPLC analysis with 2. The radiochemical yield was
5.62 (m, 5H), 5.69–5.75 (m, 1H), 5.86 and 5.88 (each t, 49%, and the radiochemical purity was 95%.
J. Label Compd. Radiopharm 2013, 56 562–572
Copyright © 2013 John Wiley & Sons, Ltd.
www.jlcr.org

K. Arimitsu et al.
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Acknowledgments
This work was supported in part by a Grant-in-Aid for General
Scientific Research from the Japan Society for the Promotion of
Science.
[11] T. Mukai, M. Hagimori, K. Arimitsu, T. Katoh, M. Ukon, T. Kajimoto, H.
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Conflict of Interest
The authors did not report any conflict of interest.
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J. Label Compd. Radiopharm 2013, 56 562–572