An unusual Wittig reaction with sugar derivatives: Exclusive formation of a 4-deoxy analogue of α-galactosyl ceramide

Article abstract of DOI:10.1039/c4ra03369h

The Wittig reaction of the pyrano-type reducing sugars undergoes an unexpected formation of dienes through the elimination of a benzyloxy group in the presence of t-BuOK. LiHMDS is used rather than t-BuOK to prevent alcohol elimination in the same sugar derivatives. Collectively, t-BuOK has unusual functions in the Wittig reaction that correspond with other bases such as LiHMDS, NaH, and n-BuLi. This unusual function of t-BuOK showed that a unique 4-deoxy-5-hydroxyl analogue 2 of α-galactosyl ceramide was formed exclusively.

Full text of DOI:10.1039/c4ra03369h

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An unusual Wittig reaction with sugar derivatives:
exclusive formation of a 4-deoxy analogue of
a-galactosyl ceramide†
Cite this: RSC Adv., 2014, 4, 26524
Ratnnadeep C. Sawant, Yu-Hsuan Lih, Shih-An Yang, Chun-Hong Yeh, Hung-Ju Tai,
*
Chung-Li Huang, Hua-Shuan Lin, Satpal Singh Badsara and Shun-Yuan Luo
The Wittig reaction of the pyrano-type reducing sugars undergoes an unexpected formation of dienes
through the elimination of a benzyloxy group in the presence of t-BuOK. LiHMDS is used rather than t-
BuOK to prevent alcohol elimination in the same sugar derivatives. Collectively, t-BuOK has unusual
functions in the Wittig reaction that correspond with other bases such as LiHMDS, NaH, and n-BuLi. This
unusual function of t-BuOK showed that a unique 4-deoxy-5-hydroxyl analogue 2 of a-galactosyl
ceramide was formed exclusively.
Received 14th April 2014
Accepted 3rd June 2014
DOI: 10.1039/c4ra03369h
www.rsc.org/advances
Introduction
The Wittig reaction is one of the most commonly used methods
for the synthesis of alkenes which has attracted substantial
interest from the scientic community.1 The mild reaction
conditions, high yields, and the absence of migration for the
formed bond are notable features of the Wittig reaction. The
remarkable success of the Wittig reaction in synthetic organic
chemistry originates from using phosphonium ylides with the
formation of phosphonium oxide as a side product, which
drives the reaction to completion. The chemistry of the Wittig
reaction was initially applied to sugars by Kuhn and Brossmer
in 1962, who reacted glyceraldehyde with (carbethoxy-
methylene) triphenyl-phosphorane.2 Thereaer, notable appli-
cations of this reaction were expanded in various directions for
approximately 6 decades.3 Usually the carbonyl olen product is
oen achieved in normal condition of Wittig reaction. Inter-
estingly, previous reports have demonstrated that 2-deoxy
sugars undergoes base promoted b-elimination to form corre-
sponding dienes.4 It is worth mentioning here that there are few
reports appeared recently describing the elimination reaction of
pyrano-sugar derivatives5 as well as a,b-elimination of anomeric
form of pyranoses which provided the dienes under the inu-
ence of base6 during the Wittig reaction.
Fig. 1 a-Galactosyl ceramide 1 and its 4-deoxy-5-hydroxyl analogue 2.
an a-linked D-galacto-pyranoside with phytosphingosine-like
ceramide. The hydroxyl groups of a-galactosyl ceramide
(a-GalCer) 1 play signicant role through hydrogen bonding
with various proteins9 that can be recognize with the help of
X-ray crystallographic analysis.20
Kim et al. disclosed the role of the 4-hydroxyl group for
CD1d-mediated NKT cell activation.10 Van Calenbergh con-
ducted synthesis and in vitro evaluation of a-GalCer epimers.11
Because of its unique structure, no study has reported a method
for the synthesis of analogue 2. However, we herein report the
straightforward synthesis of 4-deoxy-5-hydroxyl analogue 2
(Fig. 1) by applying an unusual Wittig reaction with reducing
disaccharides.
The a-galactosyl ceramide 1 (ref. 7) (a-GalCer) is also referred
to as KRN7000 (Fig. 1), and is a commonly known signicant
compound that is best characterized as an antigen for CD1d-
reactive T cells in mice and humans.8 The structure of the
4-deoxy-5-hydroxyl analog 2 (Fig. 1) of a-GalCer is composed of
Results and discussion
We recently published a concise synthesis of a-galactosyl
ceramide from D-galactosyl iodide and D-lyxose12a and con-
ducted a study to prepare the derivatives of a-galactosyl
ceramide.12b However, we attempted to prepare a-GalCer
analogues in the phytosphingolipid chain that contain one
additional hydroxyl group compared to phytosphingosine. Kim
and co-workers10 reported the 3-deoxy and 4-deoxy analogues of
a-GalCer with results of their biological evaluation but they
Department of Chemistry, National Chung Hsing University, Taichung 402, Taiwan.
E-mail: syluo@dragon.nchu.edu.tw; Fax: +886 (4)22862547
† Electronic supplementary information (ESI) available. See DOI:
10.1039/c4ra03369h
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Table 1 Effect of various bases on Wittig reaction of disaccharides
required additional steps to nalize the synthesis of related
analogues by using different precursors (Fig. 2).
We began by stereoselectively and regioselectively preparing
the key disaccharides (5a–5c) from a well-azeotroped solution of
2,3,4,6-tetra-O-benzyl-D-galactosyl iodide (3) and hexopyranoses
(4a–4c).12b The galactosyl iodide 3 (ref. 13) was prepared in situ
via the treatment of 2,3,4,6-tetra-O-benzyl-D-galactosyl acetate
with iodo-trimethylsilane. The in situ generated 3 was then
treated with hexopyranoses (4a–4c) in the presence of N,N-dii-
sopropylethylamine (DIPEA, 1 equiv.) and tetra-n-butylammo-
nium iodide (TBAI, 3 equiv.) in toluene for 1 h at 65 ꢀC, followed
by azeotropic distillation, provided a-linked disaccharides (5a–
5c) containing protected D-galactose 4a, D-glucose 4b, and D-
mannose 4c (ref. 14) (Table 1) in good overall yields.13
The Wittig reaction was performed using the in situ genera-
tion of phosphorane15 with addition of t-BuOK16 to the well-
stirred suspension of D-galactose hemiacetal 5a and phospho-
nium salt at 0 ꢀC, which produced the undesired product 6a in
3 h in an excellent yield (96%, entry 1, Table 1). The effects of
changing bases for the formation of eliminated (6a, 6b) and
expected (7a–7c) compounds during the Wittig reaction are
shown in Table 1.
Entry
SM
Base
Product (yield)
We initially expected to obtain the olenation product 7a
from the Wittig reaction of the hemiacetal of D-galactose 5a by
using t-BuOK, but 1H NMR analysis of the reaction product
indicated that one set of benzyl protons disappeared, and one
additional double bond proton appeared in the olen region of
the proton spectrum. Moreover, the 13C NMR spectrum showed
one additional downeld peak at d 154 ppm, which belongs to
diene C-5 of the sphingosine chain of 6a. The formation of 6a
and 6b are probably due to initial base-promoted elimination in
the ring-opened isomers of 5a to 5c and furnished the olena-
tion products 6a and 6b. The change in the number of equiva-
1
2
3
4
5
6
7
8
9
5a
5a
5a
5a
5a
5b
5b
5c
5c
4 equiv. t-BuOK
3 equiv. t-BuOK
3 equiv. NaH
2.5 equiv. n-BuLi
6 equiv. LiHMDS
4 equiv. t-BuOK
6 equiv. LiHMDS
4 equiv. t-BuOK
6 equiv. LiHMDS
6a (96%)
6a (56%)
7a (04%)
7a (trace)
7a (90%)
6b (66%)
7b (72%)
6b (84%)
7c (72%)
lent of t-BuOK from 4 to 3 in the similar reaction resulted in a results were observed during the Wittig reaction of the hemi-
decreased yield of product to 56% (entry 2, Table 1). Using acetal of D-glucose 5b and D-mannose 5c when using the base
sodium hydride formed only 4% of the desired product 7a (entry LiHMDS, wherein we formed the expected single olenation
3, Table 1), and n-butyl lithium resulted in trace amounts of the compounds 7b and 7c in favorable yields (entry 7 and entry 9,
desired single olenation compound 7a on the TLC plate (entry Table 1). Moreover, using t-BuOK in the Wittig reactions of the
4, Table 1). It is worth mentioning here that when we have hemiacetal of D-glucose 5b and D-mannose 5c produced the
employed some stronger bases, such as NaH16 and n-BuLi,17 we same unexpected dienes 6b in moderate to favorable yields
didn't obtain the unexpected diene derivative. However, the (entry 6 and entry 8, Table 1). Conversely, this unexpected
expected olenation products were obtained in very low yields formation of 4-deoxy sugars occurred through the elimination
(Table 1).
of the benzyloxy group at the C-3 position during the Wittig
We then applied the similar Wittig reaction exhibiting the D- reaction of sugar derivatives (5a–5c).
galactosyl hemiacetal 5a by using the hindered strong base
For further study of these unusual Wittig products, we have
LiHMDS.18 We obtained the expected olenation compound 7a, carried out the Wittig reaction of hexapyranose sugars 8 as
in an excellent yield (90%, entry 5, Table 1). This Wittig reaction shown in Table 2. In this regard, rst, we have treated galactosyl
of hemiacetal 5a with LiHMDS takes 24 hours, because the hemiacetal 8a with t-BuOK and phosphonium salt in THF at
hemiacetal cannot consume before 24 hours. The Similar 0 ꢀC (entry 1, Table 2) which provided eliminated compound 9a
in 71% isolated yield. Similarly, the reaction of glucosyl hemi-
acetal 8b (entry 3, Table 2) and mannosyl hemiacetal 8c (entry 5,
Table 2) with phosphonium salt and t-BuOK in THF at 0 ꢀC
produced eliminated compound 9b in 90 and 79% yield
respectively. We have also employed talosyl pyrano-reducing
sugar 8d for such kind of transformation under same reaction
conditions, which provided eliminated diene 9a and expected
olen 10d in good yields (entry 7 and entry 8, Table 2).
Fig. 2 a-GalCer analogues reported by Kim et al.10
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Table 2 Effect of various bases on Wittig reaction of hexopyranoses
be the real reaction partner for the Wittig reaction with phos-
phonium salt, could provide the unexpected diene 6a according
to pathway b.4a
We decided to synthesize a-GalCer derivative with diene 6a
because of their structural similarities from positions C-2 to C-4
in the a-GalCer 1 and analogue 6a (Scheme 2). We applied
Mitsunobu conditions19 to obtain the azido displacement
product 12 at the C-2 position, which successfully produced an
excellent 96% yield. Because the olen was reacted with the
azido group during intramolecular cycloaddition, we puried
azide 12 without data collection and performed the next step
directly. The azido compound 12 was used rst to form an
amine by using the Staudinger reaction followed by an amide
bond formation to produce the compound 13 in an 80% yield.
Finally, global deprotection obtained an 85% crude yield of the
nal 4-deoxy-5-hydroxyl analogue 2 by using a palladium cata-
lyst in a chloroform and methanol mixture of solvent at room
temperature. At last, the crude product was puried by per-
forming ash column chromatography on silica gel to afford
compound 2 in a 62% yield.
Entry
SM
Base
Product (yield)
1
2
3
4
5
6
7
8
8a
8a
8b
8b
8c
8c
8d
8d
4 equiv. t-BuOK
6 equiv. LiHMDS
4 equiv. t-BuOK
6 equiv. LiHMDS
4 equiv. t-BuOK
6 equiv. LiHMDS
4 equiv. t-BuOK
6 equiv. LiHMDS
9a (71%)
10a (65%)
9b (90%)
10b (72%)
9b (79%)
10c (64%)
9a (80%)
10d (87%)
The SAR studies on the crystal structure of a-GalCer com-
plexed with CD1d reveals that the analogue lacking the
4-hydroxyl group on the phytosphingosine exhibits slightly
reduced activity as compared to a-GalCer.20 However, the
activity of analogue 2 lacking the 4-hydroxyl group, which
contains an additional hydroxyl group at the C-5 position of the
We have also proposed a plausible mechanism for these phytosphingosine chain, remains worthy of investigation.
interesting unexpected transformations as shown in Scheme 1. Examining the binding activity of these newly formed
First, hemiacetal 5a underwent Wittig reaction with phospho- compounds with CD1d molecules to stimulate NKT cells would
nium ylide to provide the olen 7a via oxa-phosphetane inter- be interesting. Many studies have reported on the biological
mediate 11b (pathway a). We assumed that olen 7a can further activities of numerous derivatives with different functional
undergo b-elimination to provide the diene 6a. In this regard, groups at various positions of a-GalCer.20 Based on computer
we have treated olen 7a with 4 equiv. of t-BuOK and phos- modeling, Henon et al. suggested that the 4-OH group in the
phonium salt in THF but we could not obtained any product as sphingosine chain of a-GalCer could form hydrogen bond with
the olen 7a was intact. Therefore, we have proposed alternative Asp80 of CD1d molecule and was important for the recognition
scenario where the aldehyde form of 5a can be transformed into of mouse NKT cells.21 However, whether the 4-OH and 5-OH-
a,b-unsaturated aldehyde 11c in presence of t-BuOK, which will bonds play a role to anchor the ligand into the binding groove of
CD1d molecule is unclear. Moreover, removing the 3-OH and
2-OH groups on the sphingosine chain of a-GalCer caused no
response of splenocytes when stimulated with glycolipid-loaded
dendritic cells.22 These data suggested that the hydroxyl group
on the sphingosine chain of a-GalCer is very important.
Scheme 1 Plausible mechanism of unusual Wittig reaction.
26526 | RSC Adv., 2014, 4, 26524–26534
Scheme 2 Preparation of a-galactosyl ceramide analogue 2.
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toluene and N,N-dimethylformamide were puried and dried
from a safe purication system containing activated Al2O3. All
reagents obtained from commercial sources were used without
purication, unless otherwise mentioned. Flash column chro-
matography was carried out on Silica Gel 60 (230–400 mesh,
E. Merck). TLC was performed on pre-coated glass plates of
Silica Gel 60 F254 (0.25 mm, E. Merck); detection was executed
by spraying with
a solution of Ce(NH4)2(NO3)6 (0.5 g),
(NH4)6Mo7O24 (24 g) and H2SO4 (28 mL) in water (500 mL) and
subsequent heating on a hot plate. Optical rotations were
measured at 589 nm (Na) at ꢂ27 ꢀC. 1H, 13C NMR, DEPT, 1H–1H
COSY, H–13C COSY, and H–1H NOESY spectra were recorded
with 400 and 600 MHz instruments. Chemical shis are in ppm
from Me4Si, generated from the CDCl3 lock signal at d 7.24 ppm.
IR spectra were taken with a FT-IR spectrometer using KBr
plates. Mass spectra were analyzed on a Finnigan LTQ-Orbi-
trapxL instrument with an ESI source.
2,3,4-Tri-O-benzyl-6-O-(2,3,4,6-tetra-O-benzyl-a-D-galactopyra-
nosyl)-D-galactopyranose (5a). To a solution of 1-O-acetyl-
2,3,4,6-tetra-O-benzyl-b-D-galactopyranoside 3 (760 mg, 1.31
mmol) in anhydrous dichloromethane (8 mL) was added iodo-
trimethylsilane (232 mL, 1.63 mmol) at 0 ꢀC under nitrogen
atmosphere. Aer the reaction was stirred for 30 min, the
mixture was evaporated in vacuo. Toluene (7 mL) was added to
the residue and evaporated in vacuo for three times. In another
round bottom ask, a mixture of acceptor 4a (600 mg, 1.33
mmol), diisopropylethylamine (227 mL, 1.31 mmol), tetrabuty-
1
1
Fig. 3 Biological activity of a-GalCer analogue 2. The indicated
glycolipids (5 mM and 25 mM) were loaded onto A20-mCD1d cells and
co-cultured with mNK1.2 cells. Three days after incubation, superna-
tants were harvested to determine the production of IL-2 by ELISA.
Data were presented as means ꢁ SD. * p < 0.05 vs. 25 mM of a-GalCer
by one-way ANOVA with Tukey's multiple comparison test.
To evaluate the biological activities of a-GalCer and analog 2
(Fig. 3), B lymphoma cells overexpressing mouse CD1d/A20-
mCD1d was loaded with these glycolipids to stimulate the Va14-
expressing mNK1.2 cells to produce IL-2 and the level of IL-2
was determined by ELISA. As shown in Fig. 2, a-GalCer induced
the production of IL-2 in a dose-dependent manner (5 mM:
305 ꢁ 7.3 and 25 mM: 404 ꢁ 9.7). IL-2 induction by analog 2 at
5 mM (323.4 ꢁ 27.4) was comparable to a-GalCer, but signi-
cantly lower than a-GalCer at 25 mM (322.9 ꢁ 32.3, p < 0.05 by
one-way ANOVA with Tukey's multiple comparison test).
These results suggested that the hydroxyl group at the 4th
position of the sphingosine chain of a-GalCer is important for
the CD1d a-GalCer complex to stimulate the NKT cells. Whether
such modication at the sphingosine chain of a-GalCer could
inuence the Th1/Th2 polarization awaits future investigation.
˚
lammonium iodide (1.44 g, 3.92 mmol) and 4 A molecular
sieves in anhydrous toluene (7 mL) was stirred for 10 min at
65 ꢀC under nitrogen atmosphere. A solution of iodo-residue in
toluene (7 mL) was transferred into the reaction ask which
contains acceptor, the mixture was kept stirring for 1 h, and
ethyl acetate (20 mL) was added to the reaction ask to remove
white precipitate and molecular sieves by ltration through
celite. The resulting mixture was extracted with aqueous
Na2S2O3 (3 ꢃ 15 mL) and brine (15 mL), and the organic layers
were dried over anhydrous MgSO4, ltered, and concentrated in
vacuo. The residue was puried by column chromatography on
Conclusion
In conclusion, the Wittig reaction can be applied to extend the silica gel to afford the desired product 5a (960 mg, 75%). Rf 0.5
sphingosine chain of a-GalCer. The unexpected formation of a (EtOAc/Hex ¼ 1/2); [a]2D9 +38.9 (c 1.2, CHCl3); IR (CHCl3) n 3445,
diene analogue enables preparing unique type of a-GalCer 2 3030, 1454, 1098 cmꢄ1; a-form: 1H NMR (400 MHz, CDCl3) d
derivative. Using the base LiHMDS rather than t-BuOK prevents 7.39–7.23 (m, 35H, ArH), 5.18 (d, J ¼ 2.4 Hz, 1H, H-1), 4.91 (d, J ¼
the formation of unexpected olenation compounds. Other- 10.4 Hz, 1H, CH2Ph), 4.90 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.89 (d,
wise, elimination occurs, and diene is the only product in J ¼ 10.4 Hz, 1H, CH2Ph), 4.88 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.78
various pyrano-type reducing sugar derivatives. The mechanism (d, J ¼ 11.6 Hz, 2H, CH2Ph), 4.75 (d, J ¼ 3.2 Hz, 1H, H-10), 4.60
in which t-BuOK inuences the outcome of the reaction has (d, J ¼ 10.4 Hz, 2H, CH2Ph), 4.53 (d, J ¼ 11.6 Hz, 2H, CH2Ph),
been proposed. This unusual effect of t-BuOK has been 4.46 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.42 (d, J ¼ 12.0 Hz, 1H,
extended to the hexopyranoses of various sugar derivatives. The CH2Ph), 4.35 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.30 (d, J ¼ 11.6 Hz,
hydroxyl group at 4th position of the sphingosine chain in a- 1H, CH2Ph), 4.13 (m, 1H, H-30), 4.03–3.89 (m, 7H, H-2, H-20, H-3,
GalCer plays an important role for the interaction between H-4, H-40, H-5, H-50), 3.62–3.48 (m, 4H, H-6a, H-6b, H-6a0, H-6b0),
CD1d–glycolipid complex and T-cell receptor of NKT cells.
3.41 (s, 1H, OH); 13C NMR (150 MHz, CDCl3) d 138.62 (C), 138.58
(C), 138.5 (C), 138.4 (C ꢃ 2), 138.2 (C), 137.5 (C), 128.4 (CH ꢃ 2),
128.3 (CH ꢃ 4), 128.23 (CH ꢃ 2), 128.19 (CH ꢃ 2), 128.15 (CH ꢃ
4), 128.1 (CH ꢃ 2), 128.04 (CH ꢃ 2), 128.01 (CH ꢃ 2), 127.96
(CH ꢃ 2), 127.9 (CH ꢃ 2), 127.8 (CH), 127.61 (CH), 127.57 (CH),
Experimental section
General information
Some reactions were conducted in ame-dried glassware, under 127.5 (CH ꢃ 2), 127.4 (CH ꢃ 2), 127.34 (CH ꢃ 2), 127.30 (CH ꢃ
nitrogen atmosphere. Dichloromethane, tetrahydrofuran, 2), 98.4 (CH), 91.5 (CH), 78.9 (CH), 78.6 (CH), 76.4 (CH), 76.3
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(CH), 75.2 (CH), 74.6 (CH), 74.4 (CH2), 73.5 (CH2), 73.43 (CH2), NMR (400 MHz, CDCl3) d 7.37–7.24 (m, 35H, ArH), 5.00 (d, J ¼
73.39 (CH2), 73.3 (CH2), 73.0 (CH2), 72.9 (CH2), 72.8 (CH2), 69.7 3.6 Hz, 1H, H-10), 4.97–4.53 (m, 13H, CH2Ph, H-1), 4.44 (d, J ¼
(CH), 69.3 (CH), 68.9 (CH2); b-form: 1H NMR (400 MHz, CDCl3) d 11.2 Hz, 1H, CH2Ph), 4.30 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.07–3.39
7.37–7.23 (m, 35H, ArH), 4.82–4.63 (m, 15H, H-10, CH2Ph), 4.56 (m, 11H, H-20, H-3, H-30, H-4, H-40, H-5, H-50, H-6a, H-6b, H-6a0,
(d, J ¼ 8.4 Hz, 1H, H-1), 4.03–3.89 (m, 4H, H-20, H-3, H-30, H-40), H-6b0), 3.27 (dd, J ¼ 8.4, 8.0 Hz, 1H, H-2); 13C NMR (150 MHz,
3.82–3.68 (m, 4H, H-2, H-5, H-6a, H-6a0), 3.64–3.48 (m, 4H, H-4, CDCl3) d 138.64 (C), 138.59 (C), 138.5 (C), 138.4 (C), 138.2 (C),
H-50, H-6b, H-6b0), 3.41 (s, 1H, OH); 13C NMR (150 MHz, CDCl3) 138.1 (C), 137.5 (C), 128.3 (CH ꢃ 2), 128.24 (CH ꢃ 2), 128.18 (CH
d 138.62 (C), 138.58 (C), 138.5 (C), 138.4 (C ꢃ 2), 138.2 (C), 137.5 ꢃ 2), 128.11 (CH ꢃ 2), 128.09 (CH ꢃ 2), 128.07 (CH ꢃ 2), 128.0
(C), 128.4 (CH ꢃ 2), 128.3 (CH ꢃ 4), 128.23 (CH ꢃ 2), 128.19 (CH (CH), 127.82 (CH ꢃ 2), 127.78 (CH ꢃ 2), 127.74 (CH ꢃ 2), 127.72
ꢃ 2), 128.15 (CH ꢃ 4), 128.1 (CH ꢃ 2), 128.04 (CH ꢃ 2), 128.01 (CH ꢃ 2), 127.71 (CH ꢃ 2), 127.69 (CH ꢃ 2), 127.67 (CH ꢃ 2),
(CH ꢃ 2), 127.96 (CH ꢃ 2), 127.9 (CH ꢃ 2), 127.8 (CH), 127.61 127.65 (CH ꢃ 2), 127.6 (CH), 127.54 (CH), 127.49 (CH), 127.42
(CH), 127.57 (CH), 127.5 (CH ꢃ 2), 127.4 (CH ꢃ 2), 127.34 (CH ꢃ (CH), 127.37 (CH), 127.31 (CH), 98.1 (CH), 97.1 (CH), 84.3 (CH),
2), 127.30 (CH ꢃ 2), 98.2 (CH), 97.6 (CH), 81.9 (CH), 80.6 (CH), 83.2 (CH), 78.4 (CH), 77.6 (CH), 75.3 (CH2), 75.2 (CH), 74.8 (CH),
79.0 (CH), 76.2 (CH), 74.8 (CH2), 74.3 (CH), 74.3 (CH), 74.0 (CH), 74.7 (CH2), 74.5 (CH), 74.3 (CH2), 73.3 (CH2), 73.0 (CH2), 72.7
73.5 (CH2), 73.43 (CH2), 73.39 (CH2), 73.3 (CH2), 73.0 (CH2), 72.9 (CH2), 71.9 (CH2), 71.6 (CH), 69 (CH2), 67.9 (CH2); HRMS (ESI,
(CH2), 72.8 (CH2), 69.4 (CH), 68.4 (CH2); HRMS (ESI, M + Na+) M + Na+) calcd for C61H64O11Na 995.4341, found 995.4343.
calcd for C61H64O11Na 995.4341, found 995.4380.
2,3,4-Tri-O-bezyl-6-O-(2,3,4,6-tetra-O-benzyl-a-D-galactopyran-
2,3,4-Tri-O-benzyl-6-O-(2,3,4,6-tetra-O-benzyl-a-D-galactopyra- osyl)-D-mannopyranoside (5c). To a solution of 1-O-acetyl-
nosyl)-D-glucopyranose (5b). To a solution of 1-O-acetyl-2,3,4,6- 2,3,4,6-tetra-O-benzyl-b-D-galactopyranoside 3 (200 mg, 0.34
tetra-O-benzyl-b-D-galactopyranoside 3 (1.90 g, 3.26 mmol) in mmol) in anhydrous dichloromethane (2 mL) was added iodo-
anhydrous dichloromethane (20 mL) was added iodo- trimethylsilane (62 mL, 0.43 mmol) at 0 ꢀC under nitrogen
trimethylsilane (578 mL, 4.08 mmol) at 0 ꢀC under nitrogen atmosphere. Aer the reaction was stirred for 30 min, the
atmosphere. Aer the reaction was stirred for 30 min, the mixture was evaporated in vacuo. Toluene (2 mL) was added to
mixture was evaporated in vacuo. Toluene (17 mL) was added to the residue and evaporated in vacuo for three times. In another
the residue and evaporated in vacuo for three times. In another round bottom ask, a mixture of acceptor 4c (170 mg, 0.38
round bottom ask, a mixture of acceptor 4b (1.50 g, 3.33 mmol), diisopropylethylamine (60 mL, 0.34 mmol), tetrabuty-
˚
mmol), diisopropylethylamine (567 mL, 3.26 mmol), tetrabutyl- lammonium iodide (380 mg, 1.02 mmol) and 4 A molecular
˚
ammonium iodide (3.60 g, 9.78 mmol) and 4 A molecular sieves sieves in anhydrous toluene (2 mL) was stirred for 10 min at 65
ꢀ
in anhydrous toluene (17 mL) was stirred for 10 min at 65 C ꢀC under nitrogen atmosphere. A solution of iodo-residue in
under nitrogen atmosphere. A solution of iodo-residue in toluene (2 mL) was transferred into the reaction ask which
toluene (17 mL) was transferred into the reaction ask which contains acceptor, the mixture was kept stirring for 1 h, and
contains acceptor, the mixture was kept stirring for 1 h, and ethyl acetate (10 mL) was added to the reaction ask to remove
ethyl acetate (30 mL) was added to the reaction ask to remove white precipitate and molecular sieves by ltration through
white precipitate and molecular sieves by ltration through celite. The resulting mixture was extracted with aqueous
celite. The resulting mixture was extracted with aqueous Na2S2O3 (3 ꢃ 5 mL) and brine (5 mL), and the organic layers
Na2S2O3 (3 ꢃ 20 mL) and brine (20 mL), and the organic layers were dried over anhydrous MgSO4, ltered, and concentrated in
were dried over anhydrous MgSO4, ltered, and concentrated in vacuo. The residue was puried by column chromatography on
vacuo. The residue was puried by column chromatography on silica gel to afford the desired product 5c (229 mg, 69%). Rf 0.5
silica gel to afford the desired product 5b (2.33 g, 72%). Rf 0.5 (EtOAc/Hex ¼ 1/2); [a]2D9 +36.7 (c 0.9, CHCl3); IR (CHCl3) n 3422,
1
(EtOAc/Hex ¼ 1/2); [a]2D9 +39.0 (c 1.3, CHCl3); IR (CHCl3) n 3431, 3030, 1454, 1097 cmꢄ1; H NMR (400 MHz, CDCl3) d 7.37–7.22
3030, 1454, 1096 cmꢄ1; a-form: 1H NMR (400 MHz, CDCl3) d (m, 35H, ArH), 5.13 (s, 1H, H-1), 5.01 (d, J ¼ 3.2 Hz, 1H, H-10),
7.37–7.24 (m, 35H, ArH), 5.09 (d, J ¼ 2.8 Hz, 1H, H-1), 4.97 (d, J ¼ 4.92 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.86 (d, J ¼ 10.8 Hz, 1H,
3.2 Hz, 1H, H-10), 4.95–4.53 (m, 12H, CH2Ph), 4.42 (d, J ¼ 12.0 CH2Ph), 4.79 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.74–4.59 (m, 7H,
Hz, 1H, CH2Ph), 4.35 (d, J ¼ 12.4 Hz, 1H, CH2Ph), 4.07–4.00 (m, CH2Ph), 4.55 (d, J ¼ 10.8 Hz, 1H, CH2Ph), 4.53 (d, J ¼ 11.6 Hz,
2H, H-20, H-5), 3.97–3.83 (m, 4H, H-3, H-30, H-40, H-6a0), 3.75– 1H, CH2Ph), 4.42 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.35 (d, J ¼ 12.0
3.68 (m, 1H, H-6b0), 3.54 (m, 4H, H-4, H-50, H-6a, H-6b), 3.40 (dd, Hz, 1H, CH2Ph), 4.07–4.00 (m, 3H, H-20, H-4, H-50), 3.96–3.93
J ¼ 9.2, 3.6 Hz, 1H, H-2); 13C NMR (150 MHz, CDCl3) d 138.64 (m, 2H, H-3, H-6a), 3.87 (m, 2H, H-30, H-40), 3.78–3.71 (m, 3H, H-
(C), 138.59 (C), 138.5 (C), 138.4 (C), 138.1 (C), 137.9 (C), 137.7 2, H-5, H-6b), 3.53 (dd, J ¼ 9.6, 6.4 Hz, 1H, H-6a0), 3.42 (dd, J ¼
(C), 128.3 (CH ꢃ 2), 128.24 (CH ꢃ 2), 128.18 (CH ꢃ 2), 128.11 9.6, 6.4 Hz, 1H, H-6b0); 13C NMR (100 MHz, CDCl3) d 138.8 (C),
(CH ꢃ 2), 128.09 (CH ꢃ 2), 128.07 (CH ꢃ 2), 128.0 (CH), 127.82 138.6 (C), 138.51 (C), 137.46 (C), 138.4 (C), 138.3 (C), 137.6 (C),
(CH ꢃ 2), 127.78 (CH ꢃ 2), 127.74 (CH ꢃ 2), 127.72 (CH ꢃ 2), 128.5 (CH ꢃ 2), 128.43 (CH ꢃ 2), 128.36 (CH ꢃ 5), 128.3 (CH ꢃ
127.71 (CH ꢃ 2), 127.69 (CH ꢃ 2), 127.67 (CH ꢃ 2), 127.65 (CH 4), 128.2 (CH ꢃ 2), 128.1 (CH ꢃ 2), 128.02 (CH ꢃ 2), 128.01
ꢃ 2), 127.6 (CH), 127.54 (CH), 127.49 (CH), 127.42 (CH), 127.37 (CH ꢃ 2), 127.94 (CH ꢃ 2), 127.88 (CH ꢃ 2), 127.8 (CH), 127.7
(CH), 127.31 (CH), 98.2 (CH), 90.7 (CH), 81.6 (CH), 80.1 (CH), (CH ꢃ 2), 127.61 (CH ꢃ 2), 127.56 (CH ꢃ 2), 127.50 (CH ꢃ 2),
78.2 (CH), 77.9 (CH), 76.5 (CH), 75.5 (CH2), 74.84 (CH2), 74.83 127.47 (CH), 98.1 (CH), 92.6 (CH), 79.9 (CH), 78.7 (CH), 76.7
(CH), 74.58 (CH2), 74.56 (CH2), 73.2 (CH2), 72.8 (CH2), 72.7 (CH), 75.4 (CH), 75.1 (CH), 75.02 (CH), 74.96 (CH2), 74.7 (CH2),
1
(CH2), 70.3 (CH), 69.3 (CH), 68.9 (CH2), 67.3 (CH2); b-form: H 73.4 (CH2), 73.2 (CH2), 73.0 (CH2), 72.8 (CH2), 72.2 (CH2), 72.0
26528 | RSC Adv., 2014, 4, 26524–26534
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(CH), 69.4 (CH), 69.3 (CH2), 68.50 (CH2); HRMS (ESI, M + Na+) 4.67 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.65 (d, J ¼ 10.2 Hz, 1H,
calcd for C61H64O11Na 995.4341, found 995.4331.
CH2Ph), 4.55 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.46–4.42 (m, 3H,
(2S,3S,4Z,6Z)-3,5-di-O-benzyl-1-O-(2,3,4,6-tetra-O-benzyl-a-D- CH2Ph, H-3), 4.37 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.21 (d, J ¼ 12.0
galactopyranosyl)-octadec-4,6-dien-1,2,3,5-tetraol (6a).
A
Hz, 1H, CH2Ph), 4.06–4.01 (m, 2H, H-20, H-50), 3.93–3.85 (m, 3H,
mixture of disaccharide 5a (980 mg, 1.00 mmol), tridecanyl- H-2, H-30, H-40), 3.70–3.62 (m, 2H, H-1a, H-1b), 3.49 (d, J ¼ 6.6
triphenylphosphonium bromide (2.06 g, 4.03 mmol) and Hz, 2H, H-6a0, H-6b0), 3.04 (bs, 1H, 2-OH), 2.28–2.24 (m, 2H, H-
potassium tert-butoxide (452 mg, 4.03 mmol) in anhydrous THF 8a, H-8b), 1.41–1.20 (m, 18H, CH2), 0.87 (t, J ¼ 7.2 Hz, 3H, CH3);
(10 mL) was stirred at 0 C under nitrogen. Aer the reaction 13C NMR (150 MHz, CDCl3) d 154.1 (C), 138.8 (C), 138.7 (C),
ꢀ
mixture was kept stirring for 3 h at 0 ꢀC, water (20 mL) was 138.6 (C), 138.3 (C), 137.9 (C), 137.5 (C ꢃ 1, CH ꢃ 1), 128.34 (CH
added to quench the reaction and the mixture was extracted ꢃ 2), 128.31 (CH ꢃ 3), 128.19 (CH ꢃ 2), 128.15 (CH ꢃ 3), 128.04
with EtOAc (3 ꢃ 30 mL). The combined organic layers were (CH ꢃ 2), 127.96 (CH ꢃ 2), 127.82 (CH ꢃ 2), 127.77 (CH ꢃ 2),
washed with brine, dried over anhydrous MgSO4, ltered, and 127.64 (CH ꢃ 2), 127.61 (CH ꢃ 2), 127.5 (CH ꢃ 2), 127.4 (CH ꢃ
concentrated under vacuum to give a residue. The residue was 2), 127.33 (CH ꢃ 2), 127.25 (CH ꢃ 2), 123.2 (CH), 111.1 (CH),
puried by column chromatography to give the only diene 6a 98.2 (CH), 79.1 (CH), 76.3 (CH), 74.8 (CH), 74.7 (CH2), 73.9 (CH),
(979 mg, 96%) as colorless oil. Rf 0.5 (EtOAc/Hex ¼ 1/3); [a]2D9 73.5 (CH2), 73.3 (CH2), 72.9 (CH2), 72.3 (CH), 71.1 (CH2), 70.3
+12.9 (c 1.6, CHCl3); IR (CHCl3) n 3063, 2924, 1652, 1454, 1099 (CH2), 70.0 (CH2), 69.3 (CH), 68.7 (CH2), 31.9 (CH2), 29.62 (CH2
cmꢄ1
;
1H NMR (400 MHz, CDCl3) d 7.36–7.20 (m, 30H, ArH), ꢃ 2), 29.59 (CH2 ꢃ 2), 29.55 (CH2 ꢃ 2), 29.4 (CH2), 29.3 (CH2),
5.83 (d, J ¼ 12.0 Hz, 1H, H-6), 5.76–5.70 (m, 1H, H-7), 4.90 (d, J ¼ 29.2 (CH2), 14.1 (CH3); HRMS (ESI, M + Na+) calcd for
11.6 Hz, 1H, CH2Ph), 4.80 (d, J ¼ 3.6 Hz, 1H, H-10), 4.75 (d, J ¼
12.0 Hz, 1H, CH2Ph), 4.73–4.66 (m, 5H, CH2Ph, H-4), 4.59 (d, J ¼
C
66H80O9Na 1039.5695, found 1039.5732.
(2R,3S,4R,5S)-3,4,5-Tri-O-benzyl-1-O-(2,3,4,6-tetra-O-benzyl-
12.0 Hz, 1H, CH2Ph), 4.54 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.49–4.37 a-D-galactopyranosyl)-octadec-6-en-1,2,3,4,5-pentaol (7a).
A
(m, 4H, CH2Ph, H-3), 4.20 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.06 (t, mixture of disaccharide 5a (1.00 g, 1.03 mmol) and tridecanyl-
J ¼ 6.4 Hz, 1H, H-50), 4.01 (dd, J ¼ 9.6, 3.6 Hz, 1H, H20), 3.90–3.87 triphenylphosphonium bromide (3.15 g, 6.17 mmol) in anhy-
(m, 2H, H-30, H-40), 3.74–3.69 (m, 1H, H-2), 3.66–3.62 (dd, 1H, H- drous tetrahydrofuran (10 mL) was cooled to 0 ꢀC under
1a), 3.54 (dd, J ¼ 11.2, 3.2 Hz, 1H, H-1b), 3.50 (d, J ¼ 6.8 Hz, 2H, nitrogen. A 1.0 M solution of lithium hexamethyldisilylamide in
H-6a0, H-6b0), 3.04 (d, J ¼ 4.4 Hz, 1H, 2-OH), 2.27–2.21 (m, 2H, THF (6.2 mL, 6.17 mmol) was added to the reaction mixture and
CH2), 1.40–1.23 (m, 18H, CH2), 0.87 (t, J ¼ 6.8 Hz, 3H, CH3); 13
C
the reaction solution was stirred for 24 h at 0 ꢀC. Water (20 mL)
NMR (100 MHz, CDCl3) d 154.0 (C), 138.9 (C), 138.73 (C), 138.68 was added to quench the reaction and the mixture was extracted
(C), 138.5 (C), 138.0 (C), 137.6 (CH), 137.4 (C), 128.4 (CH ꢃ 2), with EtOAc (3 ꢃ 30 mL). The combined organic layers were
128.30 (CH ꢃ 2), 128.26 (CH ꢃ 2), 128.2 (CH ꢃ 2), 128.1 (CH ꢃ washed with brine, dried over anhydrous MgSO4, ltered, and
2), 128.00 (CH ꢃ 2), 127.98 (CH ꢃ 2), 127.9 (CH ꢃ 2), 127.8 concentrated under vacuum to give a residue. The residue was
(CH ꢃ 2), 127.7 (CH ꢃ 2), 127.61 (CH ꢃ 2), 127.55 (CH ꢃ 2), puried by column chromatography to give the olen 7a (1.05 g,
127.5 (CH ꢃ 2), 127.33 (CH ꢃ 2), 127.29 (CH ꢃ 2), 123.2 (CH), 90%, cis–trans ¼ 1.2 : 1.6) as colorless oil. Rf 0.3 (EtOAc/Hex ¼
110.5 (CH), 98.1 (CH), 79.1 (CH), 76.4 (CH), 75.0 (CH), 74.7 1/4); [a]2D8 +46.8 (c 1.6, CHCl3); IR (CHCl3) n 3483, 2925, 2360,
(CH2), 73.8 (CH), 73.4 (CH2), 73.2 (CH2), 73.0 (CH2), 72.9 (CH), 1497, 1061 cmꢄ1 1H NMR (400 MHz, CDCl3) d 7.34–7.15 (m,
;
70.9 (CH2), 70.1 (CH2), 70.0 (CH2), 69.2 (CH), 68.8 (CH2), 31.9 70H), 5.72 (dt, J ¼ 15.6, 6.8 Hz, 1H), 5.58 (dt, J ¼ 11.6, 7.2 Hz,
(CH2), 29.64 (CH2), 29.63 (CH2), 29.61 (CH2), 29.60 (CH2), 29.5 1H), 5.50–5.44 (m, 2H), 4.91 (d, J ¼ 11.2 Hz, 2H), 4.83 (d, J ¼ 4.8
(CH2), 29.4 (CH2), 29.3 (CH2), 29.2 (CH2), 22.9 (CH2), 14.1 (CH3); Hz, 2H), 4.18–4.64 (m, 11H), 4.60 (d, J ¼ 12.0 Hz, 3H), 4.54 (d, J ¼
HRMS (ESI, M + Na+) calcd for C66H80O9Na 1039.5695, found 11.2 Hz, 2H), 4.47–4.44 (m, 3H), 4.41 (d, J ¼ 11.2 Hz, 3H), 4.35
1039.5717.
(d, J ¼ 11.6 Hz, 2H), 4.27 (d, J ¼ 11.6 Hz, 3H), 4.14–4.07 (m, 2H),
(2S,3R,4Z,6Z)-3,5-Di-O-benzyl-1-O-(2,3,4,6-tetra-O-benzyl-a- 4.05 (m, 5H), 3.93–3.89 (m, 4H), 3.81–3.71 (m, 6H), 3.57 (dd, J ¼
D-galactopyranosyl)-octadec-4,6-dien-1,2,3,5-tetraol (6b).
10.4, 5.6 Hz, 2H), 3.48 (d, J ¼ 6.4 Hz, 4H), 3.26–3.21 (m, 2H),
A
mixture of disaccharide 5c (522 mg, 0.54 mmol), tridecanyl- 2.09–1.95 (m, 4H), 1.38–1.95 (m, 36H), 0.88 (t, J ¼ 6.0 Hz, 6H);
triphenylphosphonium bromide (1.09 g, 2.15 mmol) and 13C NMR (100 MHz, CDCl3) d 138.8 (C ꢃ 2), 138.6 (C ꢃ 2), 138.51
potassium tert-butoxide (240 mg, 2.15 mmol) in anhydrous THF (C ꢃ 2), 138.48 (C ꢃ 2), 138.33 (C), 137.32 (C), 138.26 (C), 138.2
(5 mL) at 0 ꢀC under nitrogen was stirred. Aer the reaction (C), 137.92 (C), 137.90 (C), 136.0 (CH), 135.0 (CH), 128.36 (CH ꢃ
mixture was kept stirring for 3 h at 0 ꢀC, water (10 mL) was 5), 128.35 (CH ꢃ 5), 128.3 (CH ꢃ 7), 128.24 (CH ꢃ 5), 128.21
added to quench the reaction and the mixture was extracted (CH ꢃ 3), 128.19 (CH ꢃ 3), 128.15 (CH ꢃ 3), 128.07 (CH ꢃ 3),
with EtOAc (3 ꢃ 20 mL). The combined organic layers were 127.06 (CH ꢃ 3), 127.9 (CH ꢃ 3), 127.8 (CH ꢃ 3), 127.7 (CH ꢃ
washed with brine, dried over anhydrous MgSO4, ltered, and 3), 127.60 (CH ꢃ 3), 127.57 (CH ꢃ 3), 127.52 (CH ꢃ 3), 127.48
concentrated under vacuum to give a residue. The residue was (CH ꢃ 3), 127.45 (CH ꢃ 3), 127.42 (CH ꢃ 3), 127.35 (CH ꢃ 3),
puried by column chromatography to give the diene 6b (457 127.3 (CH ꢃ 3), 98.12 (CH), 98.06 (CH), 82.06 (CH), 82.04 (CH),
mg, 84%) as colorless oil. Rf 0.5 (EtOAc/Hex ¼ 1/3); [a]2D4 +35.13 79.9 (CH), 79.2 (CH), 79.1 (CH), 77.40 (CH), 76.3 (CH), 75.3
(c 0.8, CHCl3); IR (CHCl3) n 3063, 2924, 1608, 1454, 1059 cmꢄ1
;
(CH2), 75.2 (CH2), 74.9 (CH ꢃ 3), 74.78 (CH2), 74.76 (CH2), 73.9
1H NMR (600 MHz, CDCl3) d 7.36–7.20 (m, 30H, ArH), 5.83 (d, (CH), 73.5 (CH2), 73.44 (CH2 ꢃ 2), 73.40 (CH2), 73.34 (CH2),
J ¼ 12.0 Hz, 1H, H-6), 5.73 (dt, J ¼ 12.0, 7.2 Hz, 1H, H-7), 4.91 (d, 73.32 (CH2), 73.0 (CH2), 72.9 (CH2), 70.4 (CH2), 70.3 (CH2), 69.80
J ¼ 11.4 Hz, 1H, CH2Ph), 4.80–4.71 (m, 6H, CH2Ph, H-10, H-4), (CH2), 69.78 (CH2), 69.5 (CH), 69.42 (CH ꢃ 2), 69.37 (CH), 68.9
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(CH2), 68.8 (CH2), 32.4 (CH2), 32.0 (CH2 ꢃ 3), 29.7 (CH2 ꢃ 5), NMR (400 MHz, CDCl3) d 7.34–7.16 (m, 35H, ArH), 5.81–5.75 (m,
29.59 (CH2), 29.55 (CH2), 29.5 (CH2), 29.4 (CH2), 29.39 (CH2), 1H, H-7), 5.45 (dd, J ¼ 11.2, 9.6 Hz, 1H, H-6), 4.91 (d, J ¼ 11.6 Hz,
29.3 (CH2), 29.2 (CH2), 28.1 (CH2), 22.7 (CH2 ꢃ 3), 14.2 (CH3 ꢃ 1H, CH2Ph), 4.87 (d, J ¼ 3.6 Hz, 1H, H-10), 4.82 (d, J ¼ 11.6 Hz,
2); HRMS (ESI, M + Na+) calcd for C73H88O10Na 1147.6270, 1H, CH2Ph), 4.73 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.67 (d, J ¼ 12.0
found 1147.6261.
Hz, 1H, CH2Ph), 4.66 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.63–4.53 (m,
(2R,3R,4R,5S,6Z)-3,4,5-Tri-O-benzyl-1-O-(2,3,4,6-tetra-O-benzyl- 6H, CH2Ph), 4.50 (t, J ¼ 9.2 Hz, 1H, H-5), 4.42 (d, J ¼ 12.0 Hz, 1H,
a-D-galactopyranosyl)-octadec-6-en-1,2,3,4,5-pentaol (7b).
CH2Ph), 4.34 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.13 (d, J ¼ 11.6 Hz,
A
mixture of disaccharide 5b (730 mg, 0.75 mmol) and trideca- 1H, CH2Ph), 4.06–4.00 (m, 4H, H-20, H-30, H-40, H-50), 3.95–3.82
nyltriphenylphosphonium bromide (2.30 g, 4.50 mmol) in (m, 2H, H-2, H-3), 3.81–3.75 (m, 3H, H-1a, H-1b, H-4), 3.51–3.44
anhydrous tetrahydrofuran (7 mL) was cooled to 0 ꢀC under (m, 2H, H-6a0, H-6b0), 2.20–2.01 (m, 2H, H-8a, H-8b), 1.40–1.25
nitrogen. A 1.0 M solution of lithium hexamethyldisilylamide in (m, 18H, CH2), 0.88 (t, J ¼ 6.4 Hz, 3H, CH3); 13C NMR (100 MHz,
THF (4.50 mL, 4.50 mmol) was added to the mixture and the CDCl3) d 138.67 (C), 138.66 (C ꢃ 2), 138.54 (C), 138.52 (C), 138.1
reaction solution was stirred for 24 h at 0 ꢀC. Water (10 mL) was (C), 137.8 (C), 136.7 (CH), 128.37 (CH ꢃ 3), 128.3 (CH ꢃ 3),
added to quench the reaction and the mixture was extracted 128.19 (CH ꢃ 6), 128.18 (CH ꢃ 4), 128.1 (CH), 128.0 (CH ꢃ 4),
with EtOAc (3 ꢃ 20 mL). The combined organic layers were 127.8 (CH), 127.70 (CH ꢃ 3), 127.65 (CH), 127.60 (CH ꢃ 3),
washed with brine, dried over anhydrous MgSO4, ltered, and 127.54 (CH), 127.48 (CH), 127.41 (CH ꢃ 2), 127.38 (CH), 127.33
concentrated under vacuum to give a residue. The residue was (CH), 127.26 (CH), 99.1 (CH), 80.6 (CH), 79.3 (CH), 78.8 (CH),
puried by column chromatography to give the olen 7b (610 76.4 (CH), 74.7 (CH2 ꢃ 1, CH ꢃ 1), 74.5 (CH2), 73.9 (CH2), 73.7
mg, 72%) as colorless oil. Rf 0.3 (EtOAc/Hex ¼ 1/4); [a]D27 +51.2 (CH2), 73.6 (CH), 73.4 (CH2), 72.9 (CH2), 71.4 (CH2), 69.73 (CH),
(c 1.1, CHCl3); IR (CHCl3) n 3463, 3030, 2925, 1497, 1061 cmꢄ1
;
69.69 (CH), 69.4 (CH2), 68.8 (CH2), 31.9 (CH2), 29.66 (CH2), 29.64
1H NMR (400 MHz, CDCl3) d 7.38–7.22 (m, 35H, ArH), 5.68–5.58 (CH2), 29.63 (CH2), 29.61 (CH2), 29.5 (CH2), 29.4 (CH2), 29.3
(m, 1H, H-7), 5.40 (t, J ¼ 10.8 Hz, 1H, H-6), 4.92 (d, J ¼ 11.4 Hz, (CH2), 28.1 (CH2), 22.7 (CH2), 14.1 (CH3); HRMS (ESI, M + Na+)
1H, CH2Ph), 4.86 (d, J ¼ 3.6 Hz, 1H, H-10), 4.82–4.51 (m, 11H, calcd for C73H88 O10Na1147.6270, found 1147.6305.
CH2Ph, H-5), 4.42 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.37 (d, J ¼ 12.0
(2R,3R,4Z,6Z)-1,3,5-Tri-O-benzyl-octadec-4,6-diene-1,2,3,5
Hz, 1H, CH2Ph), 4.34 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.04 (dd, J ¼ pentaol (9a). Method A: a solution of hemiacetal 8a (100 mg,
9.6, 3.6 Hz, 1H, H-20), 4.01–3.97 (m, 2H, H-2, H-50), 3.95–3.90 (m, 0.19 mmol) and dodecyltriphenylphosphonium bromide (378
2H, H-3, H-4), 3.81–3.68 (m, 4H, H-1a, H-1b, H-30, H-4), 3.55– mg, 0.74 mmol) in anhydrous tetrahydrofuran (1 mL) was
3.44 (m, 2H, H-6a0, H-6b0), 2.08–1.85 (m, 2H, H-8a, H-8b), 1.31– cooled to 0 ꢀC, followed by quick addition of potassium tert-
1.17 (m, 18H, CH2), 0.88 (t, J ¼ 6.8 Hz, 3H, CH3); 13C NMR (100 butoxide (83 mg, 0.74 mmol). Aer stirring for 1 h at this
MHz, CDCl3) d 138.7 (C), 138.63 (C), 138.60 (C), 138.57 (C ꢃ 2), temperature, the reaction mixture was treated with water (2 mL)
138.3 (C), 137.8 (C), 135.8 (CH), 128.4 (CH), 128.3 (CH ꢃ 5), and extracted with ethyl acetate (3 ꢃ 2 mL). The combined
128.21 (CH), 128.19 (CH ꢃ 2), 128.16 (CH ꢃ 4), 128.1 (CH ꢃ 5), organic layers were dried over anhydrous MgSO4, ltered, and
128.00 (CH), 127.98 (CH ꢃ 2), 127.9 (CH), 127.79 (CH), 127.76 concentrated. The residue was puried by column chromatog-
(CH ꢃ 2), 127.68 (CH), 127.66 (CH), 127.53 (CH), 127.49 (CH), raphy to afford olen 9a (77 mg, 71%) as a colorless oil. Method
127.44 (CH), 127.38 (CH ꢃ 3), 127.33 (CH), 127.30 (CH), 126.7 B: a solution of hemiacetal 8d (110 mg, 0.20 mmol) and dode-
(CH), 98.7 (CH), 81.8 (CH), 79.01 (CH), 78.84 (CH), 76.4 (CH), cyltriphenylphosphonium bromide (416 mg, 0.81 mmol) in
75.5 (CH), 74.9 (CH2), 74.8 (CH), 74.7 (CH2), 73.5 (CH2), 73.4 anhydrous tetrahydrofuran (1.1 mL) was cooled to 0 ꢀC, fol-
(CH2), 73.0 (CH2), 72.9 (CH2), 70.7 (CH2), 70.2 (CH2), 70.1 (CH), lowed by quick addition of potassium tert-butoxide (91 mg, 0.81
70.0 (CH), 68.8 (CH2), 31.9 (CH2), 29.7 (CH2), 29.64 (CH2), 29.63 mmol). Aer stirring for 1 h at this temperature, the reaction
(CH2), 29.61 (CH2), 29.5 (CH2), 29.43 (CH2), 29.3 (CH2), 28.2 mixture was treated with water (1.2 mL) and extracted with ethyl
(CH2), 22.7 (CH2), 14.1 (CH3); HRMS (ESI, M + Na+) calcd for acetate (3 ꢃ 2 mL). The combined organic layers were dried over
C73H88O10Na 1147.6270, found 1147.6296.
anhydrous MgSO4, ltered, and concentrated. The residue was
(2R,3R,4R,5R,6Z)-3,4,5-Tri-O-benzyl-1-O-(2,3,4,6-tetra-O-benzyl- puried by column chromatography to afford olen 9a (95 mg,
a-D-galactopyranosyl)-octadec-6-en-1,2,3,4,5-pentaol (7c).
A
80%) as a colorless oil. Rf 0.56 (EtOAc/Hex ¼ 1/3); [a]2D2 ꢄ9.1
mixture of disaccharide 5c (903 mg, 0.92 mmol) and trideca- (c 0.8, CH2Cl2); IR (CH2Cl2) n 2925, 2854, 1729, 1652, 1497, 1455
nyltriphenylphosphonium bromide (2.84 g, 5.57 mmol) in cmꢄ1 1H NMR (400 MHz, CDCl3) d 7.61–7.24 (m, 15H, ArH),
;
anhydrous tetrahydrofuran (10 mL) was cooled to 0 ꢀC under 5.86 (d, J ¼ 11.6 Hz, 1H, H-6), 5.76 (dt, J ¼ 11.6, 7.2 Hz, 1H, H-7),
nitrogen. A 1.0 M solution of lithium hexamethyldisilylamide in 7.41 (s, 2H, CH2Ph), 4.67 (d, J ¼ 9.2 Hz, 1H, H-4), 4.54 (d, J ¼ 12.0
THF (5.6 mL, 5.6 mmol) was added to the mixture and the Hz, 1H, CH2Ph), 4.49 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.44 (d, J ¼
reaction solution was stirred for 24 h at 0 ꢀC. Water (10 mL) was 11.6 Hz, 1H, CH2Ph), 4.45–4.41 (m, 1H, H-3), 4.25 (d, J ¼ 11.6
added to quench the reaction and the mixture was extracted Hz, 1H, CH2Ph), 3.72 (dddd, J ¼ 10.4, 7.2, 3.6, 3.6 Hz, 1H, H-2),
with EtOAc (3 ꢃ 20 mL). The combined organic layers were 3.50 (dd, J ¼ 10.0, 3.6 Hz, 1H, H-1a), 3.43 (dd, J ¼ 10.0, 6.8 Hz,
washed with brine, dried over anhydrous MgSO4, ltered, and 1H, H-1b), 2.78 (d, J ¼ 3.6 Hz, 1H, 2-OH), 2.26–2.20 (m, 2H,
concentrated under vacuum to give a residue. The residue was CH2), 1.40–1.24 (m, 18H, CH2), 0.88 (t, J ¼ 6.8 Hz, 3H, CH3); 13
C
puried by column chromatography to give the olen 7c (726 NMR (100 MHz, CDCl3) d 154.3 (C), 138.5 (C), 138.3 (C), 137.9
mg, 70%) as colorless oil. Rf 0.3 (EtOAc/Hex ¼ 1/4); [a]2D7 +42.6 (c (CH), 137.3 (C), 128.4 (CH ꢃ 2), 128.3 (CH ꢃ 2), 128.2 (CH ꢃ 2),
1.1, CHCl3); IR (CHCl3) n 3400, 3089, 2924, 1455, 1095 cmꢄ1; 1H 128.1 (CH ꢃ 2), 127.9 (CH), 127.8 (CH ꢃ 2), 127.7 (CH ꢃ 2),
26530 | RSC Adv., 2014, 4, 26524–26534
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127.5 (CH ꢃ 2), 123.1 (CH), 110.4 (CH), 73.8 (CH), 73.5 (CH), (m, 2H, ArH), 5.65 (dt, J ¼ 10.8, 7.2 Hz, 1H, H-7), 5.44 (dd, J ¼
73.4 (CH2), 71.2 (CH2), 71.0 (CH2), 70.1 (CH2), 31.9 (CH2), 29.65 11.4, 9.6 Hz, 1H, H-6), 4.79 (d, J ¼ 10.8 Hz, 1H, CH2Ph), 4.74 (d,
(CH2), 29.64 (CH2 ꢃ 2), 29.56 (CH2), 29.54 (CH2), 29.4 (CH2), J ¼ 10.8 Hz, 1H, CH2Ph), 4.63 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.48
29.3 (CH2), 29.2 (CH2), 22.7 (CH2), 14.1 (CH3); HRMS (ESI, M + (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.44 (dd, J ¼ 9.0, 3.6 Hz, 1H, H-5),
Na+) calcd for C39H52O4Na 607.3758, found 607.3762.
4.41 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.40 (d, J ¼ 11.4 Hz, 1H,
(2R,3S,4Z,6Z)-1,3,5-Tri-O-benzyl-octadec-4,6-diene-1,2,3,5- CH2Ph), 4.37 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.31 (d, J ¼ 12.0 Hz,
tetraol (9b). Method A: a solution of hemiacetal 8b (100 mg, 0.19 1H, CH2Ph), 4.10–4.07 (m, 1H, H-2), 3.79–3.76 (m, 2H, H-3, H-4),
mmol) and dodecyltriphenylphosphonium bromide (378 mg, 3.51 (dd, J ¼ 9.6, 6.6 Hz, 1H, H-1a), 3.47 (dd, J ¼ 9.0, 6.6 Hz, 1H,
0.74 mmol) in anhydrous tetrahydrofuran (5 mL) was cooled to H-1b), 3.13 (d, J ¼ 4.8 Hz, 1H, 2-OH), 2.06–1.95 (m, 2H, CH2),
0 ꢀC, followed by quick addition of potassium tert-butoxide (83 1.32–1.25 (m, 18H, CH2), 0.88 (t, J ¼ 7.2 Hz, 3H, CH3); 13C NMR
mg, 0.74 mmol). Aer stirring for 4 h at this temperature, the (150 MHz, CDCl3) d 138.4 (C), 138.22 (C), 138.15 (C), 138.0 (C),
reaction mixture was treated with water (2 mL) and extracted 135.5 (CH), 128.3 (CH ꢃ 8), 128.1 (CH ꢃ 2), 127.99 (CH ꢃ 2),
with ethyl acetate (3 ꢃ 2 mL). The combined organic layers were 127.97 (CH ꢃ 2), 127.7 (CH ꢃ 3), 127.6 (CH), 127.53 (CH),
dried over anhydrous MgSO4, ltered, and concentrated. The 127.50 (CH), 127.0 (CH), 82.5 (CH), 76.6 (CH), 75.4 (CH2), 74.3
residue was puried by column chromatography to afford the (CH), 73.11 (CH2), 73.06 (CH2), 71.2 (CH2), 69.9 (CH2), 69.8 (CH),
olen 9b (98 mg, 90%) as a colorless oil. Method B: a solution of 31.9 (CH2), 29.7 (CH2 ꢃ 3), 29.6 (CH2), 29.5 (CH2), 29.42 (CH2),
hemiacetal 8c (200 mg, 0.37 mmol) and dodecyltriphenyl- 29.36 (CH2), 28.1 (CH2), 22.7 (CH2), 14.1 (CH3); HRMS (ESI, M +
phosphonium bromide (760 mg, 1.48 mmol) in anhydrous Na+) calcd for C46H60O5Na 715.4333, found 715.4348. 10a-E: Rf
tetrahydrofuran (2 mL) was cooled to 0 ꢀC, followed by quick 0.44 (EtOAc/Hex ¼ 1/4); [a]2D3 ꢄ0.1 (c 0.1 CH2Cl2); IR (CH2Cl2) n
1
addition of potassium tert-butoxide (170 mg, 1.48 mmol). Aer 3361, 2925, 2853, 1649, 1497, 1209 cmꢄ1; H NMR (600 MHz,
stirring for 2 h at this temperature, the reaction mixture was CDCl3) d 7.36–7.25 (m, 18H, ArH), 7.18–7.17 (m, 2H, ArH), 5.70
treated with water (5 mL) and extracted with ethyl acetate (3 ꢃ (dt, J ¼ 15.0, 6.6 Hz, 1H, H-7), 5.40 (dd, J ¼ 15.6, 9.0 Hz, 1H, H-6),
5 mL). The combined organic layers were dried over anhydrous 4.78 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.75 (d, J ¼ 11.4 Hz, 1H,
MgSO4, ltered, and concentrated. The residue was puried by CH2Ph), 4.62 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.47 (d, J ¼ 12.0 Hz,
column chromatography to afford the olen 9b (170 mg, 79%) 1H, CH2Ph), 4.43 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.39 (d, J ¼ 12.0
as a colorless oil. Rf 0.55 (EtOAc/Hex ¼ 1/3); [a]2D3 +15.3 (c 0.5, Hz, 1H, CH2Ph), 4.35 (d, J ¼ 10.8 Hz, 1H, CH2Ph), 4.33 (d, J ¼
CH2Cl2); IR (CH2Cl2) n 3567, 3463, 3064, 3031, 2925, 2854, 1651, 11.4 Hz, 1H, CH2Ph), 4.11–4.08 (m, 1H, H-2), 4.01 (dd, J ¼ 8.4,
1496, 1457 cmꢄ1
;
1H NMR (600 MHz, CDCl3) d 7.35–7.22 (m, 5.4 Hz, 1H, H-5), 3.8 (dd, J ¼ 4.8, 4.8 Hz, 1H, H-4), 3.76–3.75 (m,
15H, ArH), 5.76 (d, J ¼ 11.4 Hz, 1H, H-6), 5.77 (dt, J ¼ 12.0, 7.8 1H, H-3), 3.51 (dd, J ¼ 9.0, 6.0 Hz, 1H, H-1a), 3.47 (dd, J ¼ 9.6, 6.6
Hz, 1H, H-7), 4.87 (d, J ¼ 9.6 Hz, 1H, H-4), 4.80 (s, 2H, CH2Ph), Hz, 1H, H-1b), 3.19 (d, J ¼ 4.8 Hz, 1H, 2-OH), 2.04–1.98 (m, 2H,
4.59 (dd, J ¼ 9.6, 4.2 Hz, 1H, H-3), 4.52–4.48 (m, 3H, CH2Ph), CH2), 1.33–1.25 (m, 18H, CH2), 0.88 (t, J ¼ 7.2 Hz, 3H, CH3); 13
C
4.29 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 3.92–3.89 (m, 1H, H-2), 3.69 (d, NMR (150 MHz, CDCl3) d 138.5 (C), 138.3 (C), 138.2 (C), 138.0
J ¼ 4.8 Hz, 1H, 2-OH), 3.56 (dd, J ¼ 9.6, 4.2 Hz, 1H, H-1a), 3.50 (C), 136.6 (CH), 128.3 (CH ꢃ 8), 128.12 (CH ꢃ 2), 128.07 (CH ꢃ
(dd, J ¼ 9.6, 6.0 Hz, 1H, H-1b), 2.35–2.31 (m, 2H, CH2), 1.42–1.40 2), 127.9 (CH ꢃ 2), 127.70 (CH), 127.66 (CH ꢃ 2), 127.62 (CH),
(m, 2H, CH2), 1.31–1.26 (m, 16H, CH2), 0.88 (t, J ¼ 7.2 Hz, 3H, 127.53 (CH), 127.46 (CH), 127.0 (CH), 82.5 (CH), 80.6 (CH), 76.3
CH3); 13C NMR (150 MHz, CDCl3) 155.0 (C), 140.3 (C), 139.8 (C), (CH), 75.4 (CH2), 73.1 (CH2), 72.8 (CH2), 71.1 (CH2), 69.9 (CH2),
138.8 (C), 137.6 (CH), 129.1 (CH ꢃ 2), 128.9 (CH ꢃ 2), 128.8 (CH 69.7 (CH), 32.4 (CH2), 31.9 (CH2), 29.70 (CH2), 29.66 (CH2 ꢃ 2),
ꢃ 2), 128.7 (CH ꢃ 2), 128.5 (CH), 128.3 (CH ꢃ 2), 128.2 (CH ꢃ 2), 29.5 (CH2), 29.4 (CH2), 29.3 (CH2), 29.0 (CH2), 22.7 (CH2), 14.1
128.0 (CH), 127.8 (CH), 124.6 (CH), 112.1 (CH), 75.3 (CH), 73.6 (CH3); HRMS (ESI, M + Na+) calcd for C46H60O5Na 715.4333,
(CH), 73.5 (CH), 72.7 (CH), 71.9 (CH2), 70.5 (CH2), 32.6 (CH2), found 715.4350.
30.34 (CH2), 30.29 (CH2), 30.16 (CH2), 30.11 (CH2), 30.0 (CH2),
29.8 (CH2), 29.4 (CH2), 23.3 (CH2), 14.3 (CH3); HRMS (ESI, M + pentaol (10b). A solution of hemiacetal 8b (100 mg, 0.19 mmol)
Na+) calcd for C39H52O4Na 607.3758, found 607.3738.
and dodecyltriphenylphosphonium bromide (567 mg, 1.11
(2S,3R,4R,5S,6Z)-1,3,4,5-Tetra-O-benzyl-octadec-6-ene-1,2,3,4,5-
(2S,3S,4R,5S,6Z)-1,3,4,5-Tetra-O-benzyl-octadec-6-ene-1,2,3,4,5- mmol) in anhydrous tetrahydrofuran (1.5 mL) was cooled to
pentaol (10a). A solution of hemiacetal 8a (100 mg, 0.19 mmol) 0 ꢀC, followed by slow addition of lithium hexamethyldisilazane
and dodecyltriphenylphosphonium bromide (567 mg, 1.11 (1 M in tetrahydrofuran, 1.11 mL, 1.11 mmol). Aer stirring for
mmol) in anhydrous tetrahydrofuran (1 mL) was cooled to 0 ꢀC, 24 h at this temperature, the reaction mixture was treated with
followed by slow addition of lithium hexamethyldisilazane (1 M water (1 mL) and extracted with ethyl acetate (3 ꢃ 2 mL). The
in tetrahydrofuran, 1.1 mL, 1.11 mmol). Aer stirring for 24 h at combined organic layers were dried over anhydrous MgSO4,
this temperature, the reaction mixture was treated with water (1 ltered, and concentrated. The residue was puried by column
mL) and extracted with ethyl acetate (3 ꢃ 2 mL). The combined chromatography to afford olen 10b (94 mg, 72%) as a colorless
organic layers were dried over anhydrous MgSO4, ltered, and oil. Rf 0.38 (EtOAc/Hex ¼ 1/6); [a]2D3 +26.5 (c 0.7, CH2Cl2); IR
concentrated. The residue was puried by column chromatog- (CH2Cl2) n 3565, 3475, 3088, 2925, 2854, 1600, 1496, 1457 cmꢄ1
;
raphy to afford the olen 10a (83 mg, 65%, Z/E ¼ 2.4/1) as a 1H NMR (600 MHz, CDCl3) d 7.37–7.19 (m, 20H, ArH), 5.67 (dt,
colorless oil. 10a-Z: Rf 0.48 (EtOAc/Hex ¼ 1/4); [a]2D2 ꢄ3.5 (c 0.1, J ¼ 10.8, 7.2 Hz, 1H, H-7), 5.47 (dd, J ¼ 11.4, 9.6 Hz, 1H, H-6),
CH2Cl2); IR (CH2Cl2) n 3468, 2925, 2854, 1648, 1496, 1210 cmꢄ1
;
4.80 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.74–4.68 (m, 2H, CH2Ph), 4.62
1H NMR (600 MHz, CDCl3) d 7.36–7.23 (m, 18H, ArH), 7.17–7.16 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.58–4.46 (m, 4H, CH2Ph, H-5), 4.37
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(d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.01 (bs, 1H, H-2), 3.77–3.72 (m, 2H, CH2Ph), 4.49–4.41 (m, 3H, CH2Ph, H-5), 4.33 (d, J ¼ 11.6 Hz, 1H,
H-4, H-3), 3.61–3.56 (m, 2H, H-1a, H-1b), 2.98 (d, J ¼ 4.2 Hz, 1H, CH2Ph), 4.17 (dddd, J ¼ 8.0, 6.0, 6.0, 2.0 Hz, 1H, H-2), 3.90 (dd,
2-OH), 2.07–2.01 (m, 2H, CH2), 1.30–1.16 (m, 18H, CH2), 0.88 (t, J ¼ 4.8, 4.8 Hz, 1H, H-4), 3.80 (dd, J ¼ 4.8, 2.0 Hz, 1H, H-3), 3.54
J ¼ 7.2 Hz, 3H, CH3); 13C NMR (150 MHz, CDCl3) d 138.45 (C), (d, J ¼ 6.0 Hz, 2H, H-1a, H-1b), 3.44 (d, J ¼ 4.4 Hz, 1H, 2-OH),
138.38 (C), 138.35 (C), 138.1 (C), 135.6 (CH), 128.55 (CH), 128.48 2.06–1.88 (m, 2H, CH2), 1.30–1.20 (m, 18H, CH2), 0.88 (t, J ¼ 7.2
(CH), 128.4 (CH), 128.3 (CH ꢃ 7), 128.2 (CH ꢃ 2), 127.9 (CH ꢃ Hz, 3H, CH3); 13C NMR (100 MHz, CDCl3) d 138.3 (C), 138.2 (C),
2), 127.8 (CH ꢃ 2), 127.7 (CH), 127.6 (CH), 127.5 (CH), 127.4 138.1 (C), 137.9 (C), 136.6 (CH), 128.3 (CH ꢃ 8), 128.1 (CH ꢃ 2),
(CH), 127.0 (CH), 126.9 (CH), 81.8 (CH), 78.3 (CH), 74.9 (CH), 127.9 (CH ꢃ 2), 127.8 (CH ꢃ 2), 127.7 (CH ꢃ 2), 127.66 (CH),
74.7 (CH2), 73.3 (CH2), 73.1 (CH2), 71.2 (CH2), 70.5 (CH), 70.2 127.61 (CH), 127.51 (CH), 127.47 (CH), 126.5 (CH), 81.9 (CH),
(CH2), 31.9 (CH2), 29.7 (CH2), 29.6 (CH2 ꢃ 2), 29.5 (CH2), 29.4 77.2 (CH), 74.4 (CH2), 74.1 (CH), 73.2 (CH2), 72.7 (CH2), 71.1
(CH2), 29.3 (CH2), 28.1 (CH2), 22.7 (CH2), 14.1 (CH3); HRMS (ESI, (CH2), 70.1 (CH), 70.0 (CH2), 31.9 (CH2), 29.7 (CH2), 29.63
M + Na+) calcd for C46H60O5Na 715.4333, found 715.4349.
(2S,3R,4R,5R)-1,3,4,5-Tetra-O-benzyl-octadec-6-ene-1,2,3,4,5- (CH2), 22.7 (CH2), 14.1 (CH3); HRMS (ESI, M + Na+) calcd for
(CH2 ꢃ 2), 29.60 (CH2), 29.5 (CH2), 29.4 (CH2), 29.3 (CH2), 28.2
pentaol (10c). A solution of hemiacetal 8c (100 mg, 0.18 mmol)
C46H60O5Na 715.4333, found 715.4331. 10d-E: Rf 0.59 (EtOAc/
and dodecyltriphenylphosphonium bromide (567 mg, 1.11 Hex ¼ 1/4); [a]2D7 ꢄ29.6 (c 1.0 CH2Cl2); IR (CH2Cl2) n 3509, 3061,
mmol) in anhydrous tetrahydrofuran (1 mL) was cooled to 0 ꢀC, 3031, 2925, 2854, 1496, 1456, 1209, 1097, 1065, 1027, 973 cmꢄ1
;
followed by slow addition of lithium hexamethyldisilazane (1 M 1H NMR (400 MHz, CDCl3) d 7.32–7.19 (m, 20H, ArH), 5.63 (dt,
in tetrahydrofuran, 1.1 mL, 1.11 mmol). Aer stirring for 12 h at J ¼ 15.6, 6.4 Hz, 1H, H-7), 5.51 (dd, J ¼ 15.6, 8.4 Hz, 1H, H-6),
this temperature, the reaction mixture was treated with water (2 4.80 (d, J ¼ 10.8 Hz, 1H, CH2Ph), 4.66 (d, J ¼ 11.2 Hz, 1H,
mL) and extracted with ethyl acetate (3 ꢃ 2 mL). The combined CH2Ph), 4.59 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.56 (d, J ¼ 10.8 Hz,
organic layers were dried over anhydrous MgSO4, ltered, and 1H, CH2Ph), 4.51 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.47 (d, J ¼ 12.0
concentrated. The residue was puried by column chromatog- Hz, 1H, CH2Ph), 4.44 (d, J ¼ 12.0 Hz, 1H, CH2Ph), 4.32 (d, J ¼
raphy to afford olen 10c (82 mg, 64%) as a colorless oil. Rf 0.68 12.0 Hz, 1H, CH2Ph), 4.16–4.11 (m, 1H, H-2), 4.00 (dd, J ¼ 8.4,
(EtOAc/Hex ¼ 1/3); [a]D23 ꢄ7.3 (c 4.8, CH2Cl2); IR (CHCl3) n 3565, 4.8 Hz, 1H, H-5), 3.91 (dd, J ¼ 5.2, 5.2 Hz, 1H, H-4), 3.73 (dd, J ¼
1
3475, 3064, 3031, 2925, 2854, 1599, 1495, 1457 cmꢄ1; H NMR 6.0, 2.0 Hz, 1H, H-3), 3.53 (d, J ¼ 6.4 Hz, 2H, H-1a, H-1b), 3.20 (d,
(600 MHz, CDCl3) d 7.38–7.18 (m, 20H, ArH), 5.83–5.77 (m, 1H, J ¼ 5.6 Hz, 1H, 2-OH), 2.11–2.06 (m, 2H, CH2), 1.40–1.26 (m,
H-7), 5.54–5.48 (m, 1H, H-6), 4.72–4.04 (m, 9H, CH2Ph, H-5), 18H, CH2), 0.88 (t, J ¼ 6.8 Hz, 3H, CH3); 13C NMR (100 MHz,
4.00 (bs, 1H, H-2), 3.89–3.56 (m, 1H, H-3), 3.84–3.83 (m, 1H, H- CDCl3) d 138.4 (C), 138.2 (C), 138.1 (C), 138.0 (C), 137.6 (CH),
4), 3.65–3.55 (m, 2H, H-1), 2.61 (bs, 1H, 2-OH), 2.17–2.05 (m, 2H, 128.31 (CH ꢃ 2), 128.29 (CH ꢃ 4), 128.27 (CH ꢃ 2), 128.1 (CH ꢃ
CH2), 1.37–1.25 (m, 18H, CH2), 0.89–0.87 (m, 3H, CH3); 13C 2), 128.0 (CH ꢃ 2), 127.8 (CH ꢃ 2), 127.7 (CH ꢃ 4), 127.5 (CH),
NMR (150 MHz, CDCl3) d 138.6 (C), 138.51 (C), 138.47 (C), 138.4 127.4 (CH), 126.5 (CH), 81.7 (CH), 80.6 (CH), 77.1 (CH), 74.3
(C), 136.7 (CH), 128.3 (CH ꢃ 8), 128.21 (CH), 128.16 (CH), 128.13 (CH2), 73.2 (CH2), 73.0 (CH2), 71.1 (CH2), 69.9 (CH), 69.8 (CH2),
(CH), 128.09 (CH), 127.8 (CH), 127.7 (CH ꢃ 2), 127.6 (CH ꢃ 2), 32.5 (CH2), 31.9 (CH2), 29.7 (CH2), 29.6 (CH2 ꢃ 2), 29.5 (CH2),
127.5 (CH), 127.44 (CH), 127.41 (CH), 127.37 (CH), 80.8 (CH), 29.35 (CH2), 29.29 (CH2), 29.20 (CH2), 22.7 (CH2), 14.1 (CH3);
79.9 (CH), 74.4 (CH2), 74.3 (CH2), 73.93 (CH2), 73.6 (CH), 73.2 HRMS (ESI, M + Na+) calcd for C46H60O5Na 715.4333, found
(CH2), 71.3 (CH2), 70.07 (CH), 69.4 (CH2), 31.9 (CH2), 29.65 715.4340.
(CH2), 29.62 (CH2), 29.60 (CH2), 29.51 (CH2), 29.47 (CH2), 29.4
(2S,3S,4Z,6Z)-2-Hexacosanoylamino-3,5-di-O-benzyl-1-O-
(CH2), 29.32 (CH2), 28.1 (CH2), 22.7 (CH2), 14.1 (CH3); HRMS (2,3,4,6-tetra-O-benzyl-a-D-galactopyranosyl)-octadec-4,6-dien-
(ESI, M + Na+) calcd for C46H60O5Na 715.4333, found 715.4343. 1,3,5-triol (13). To a solution of compound 6a (150 mg, 0.15
(2S,3S,4R,5R,6Z)-1,3,4,5-Tetra-O-benzyl-octadec-6-ene-1,2,3,4,5- mmol) and triphenylphꢀosphine (105 mg, 0.40 mmol) in anhy-
pentaol (10d). A solution of hemiacetal 8d (168 mg, 0.31 mmol) drous THF (2 mL) at 0 C was added diisopropyl azodicarbox-
and dodecyltriphenylphosphonium bromide (954 mg, 1.86 ylate (DIAD, 95 mL, 0.43 mmol) followed by dropwise addition of
mmol) in anhydrous tetrahydrofuran (1.7 mL) was cooled to diphenylphosphoryl azide (DPPA, 78 mL, 0.40 mmol) to the
0 ꢀC, followed by slow addition of lithium hexamethyldisilazane reaction ask. Aer complete addition, the temperature was
(1 M in tetrahydrofuran, 1.9 mL, 1.86 mmol). Aer stirring for brought to room temperature by removing ice bath and the
18 h at this temperature, the reaction mixture was treated with reaction was stirred for 1 h. Upon completion of the reaction,
water (2 mL) and extracted with ethyl acetate (3 ꢃ 2 mL). The the mixture was diluted with EtOAc, and the resulting mixture
combined organic layers were dried over anhydrous MgSO4, was washed by water. The organic layer was dried over anhy-
ltered, and concentrated. The residue was puried by column drous MgSO4, ltered, and concentrated in vacuo. Purication
chromatography to afford olen 10d (188 mg, 87%, E/Z ¼ 1/2) as of this residue by column chromatography gives the compound
a colorless oil. 10d-Z: Rf 0.65 (EtOAc/Hex ¼ 1/4); [a]2D6 ꢄ39.1 12 (147 mg, 97%). The compound 12 (686 mg, 0.66 mmol) was
(c 1.0, CH2Cl2); IR (CH2Cl2) n 3058, 3064, 3031, 2925, 2854, 1496, added to a round bottom ask followed by the addition of tri-
1456, 1092, 1066, 1027 cmꢄ1; 1H NMR (400 MHz, CDCl3) d 7.29– phenylphosphine (313 mg, 1.19 mmol), pyridine (2.0 mL), water
7.22 (m, 20H, ArH), 5.77 (dt, J ¼ 11.2, 7.6 Hz, 1H, H-7), 5.48 (dd, (700 mL), and tetrahydrofuran (7.0 mL). Then, the mixture was
J ¼ 10.4, 10.0 Hz, 1H, H-6), 4.73 (d, J ¼ 11.2 Hz, 1H, CH2Ph), 4.66 warmed up to 60 ꢀC and stirred for 12 h. The solvent was
(d, J ¼ 11.2 Hz, 1H, CH2Ph), 4.59 (d, J ¼ 11.6 Hz, 1H, CH2Ph), evaporated in vacuum to get a crude amine. This crude amine
4.58 (d, J ¼ 11.6 Hz, 1H, CH2Ph), 4.50 (d, J ¼ 11.6 Hz, 1H, was dissolved in anhydrous dichloromethane (7.0 mL) at room
26532 | RSC Adv., 2014, 4, 26524–26534
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13
ꢀ
temperature, 1-[3-(di-methylamino)propyl]-3-ethylcarbodiimide 0.83 (m, 6H, CH3); C NMR (150 MHz, d5-Pyridine 100 C) d
hydrochloride (EDC, 205 mg, 1.07 mmol), hexaeicosanoic acid 173.4 (C), 101.8 (CH), 73.1 (CH), 72.1 (CH2), 71.9 (CH ꢃ 2), 71.3
(306 mg, 0.77 mmol) and HOBt (145 mg, 1.07 mmol) were (CH ꢃ 2), 70.8 (CH), 63.1 (CH2), 50.0 (CH), 43.2 (CH2 ꢃ 2), 39.8
sequentially added to the solution, and the mixture was (CH2), 37.1 (CH2), 32.3 (CH2 ꢃ 4), 30.1 (CH2 ꢃ 13), 30.0 (CH2 ꢃ
continuously stirred for 12 h. The reaction solution was diluted 5), 29.9 (CH2 ꢃ 5), 26.9 (CH2), 26.4 (CH2), 24.5 (CH2), 23.0
with EtOAc, and the resulting mixture was washed by water. The (CH2 ꢃ 3), 14.2 (CH3 ꢃ 2). HRMS (ESI, M ꢄ H+) calcd for
organic layer was dried over anhydrous MgSO4, ltered, and
concentrated in vacuo. This resulting residue was puried by
column chromatography to give the amide compound 13 (730
mg, 80%). Rf 0.4 (EtOAc/Hex ¼ 1/5); [a]2D9 +10.2 (c 1.3, CHCl3);
mp ¼ 79 ꢀC; IR (CHCl3) n 3428, 2918, 2360, 1645, 1454, 1116
C50H98O9N 856.7236, found 856.7222.
General procedure for ELISA
Supernatant was collected three days aer incubation and the
production of IL-2 was quantied with DuoSet ELISA Develop-
ment System (R&D System, MN, USA). Briey, capture antibody
was coated on the plate overnight at 4 ꢀC. The plates were
washed and blocked with blocking buffer (1% BSA in PBS).
Then, samples were added and incubated for 2 hours at room
temperature, washed three times with wash buffer (0.05%
Tween 20 in PBS), added the detection antibody, and incubated
for 2 hours at room temperature. Aer washing, the plates were
added streptavidin–HRP and incubated for 20 minutes at room
temperature. Aer incubation, the plates were washed, then
added substrate solution to each well, and incubated for
20 minutes at room temperature. Finally, added stop solution
(2.0 N H2SO4) and determined the optical density (OD450) by
using a microplate reader (SpectraMax M2, Molecular device,
CA, USA).
cmꢄ1 1H NMR (600 MHz, CDCl3) d 7.37–7.19 (m, 30H, ArH),
;
5.91 (d, J ¼ 9.1 Hz, 1H, NH), 5.78 (d, J ¼ 11.8 Hz, 1H, H-6), 5.73–
5.69 (m, 1H, H-7), 4.90 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.87 (d, J ¼
3.6 Hz, 1H, H-10), 4.79 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.73 (d,
J ¼ 12.0 Hz, 3H, CH2Ph), 4.69 (d, J ¼ 9.6 Hz, 1H, H-4), 4.65 (d, J ¼
11.4 Hz, 2H, CH2Ph), 4.61 (dd, J ¼ 8.4, 9.0 Hz, 1H, H-3), 4.55 (d,
J ¼ 11.4 Hz, 1H, CH2Ph), 4.45 (d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.372
(d, J ¼ 11.4 Hz, 1H, CH2Ph), 4.367 (d, J ¼ 11.4 Hz, 1H, CH2Ph),
4.18–4.12 (m, 2H, CH2Ph, H-2), 4.04 (dd, J ¼ 9.9, 3.6 Hz, 1H, H-
20), 3.94–3.89 (m, 4H, H-1a, H-30, H-40, H-50), 3.72 (dd, J ¼ 10.8,
3.0 Hz, 1H, H-1b), 3.50–3.43 (m, 2H, H-6a0, H-6b0), 2.22–2.11 (m,
2H, H-8a, H-8b), 1.98–1.86 (m, 2H, CH2), 1.51–1.46 (m, 2H,
CH2), 1.34–1.23 (m, 62H, CH2), 0.88 (t, J ¼ 6.6 Hz, 6H, CH3 ꢃ 2);
13C NMR (150 MHz, CDCl3) d 172.6 (C), 153.7 (C), 138.8 (C),
138.7 (C), 138.6 (C), 138.5 (C), 137.8 (C), 137.7 (C), 137.5 (CH),
128.33 (CH ꢃ 4), 128.29 (CH ꢃ 4), 128.24 (CH), 128.17 (CH ꢃ 3),
127.7 (CH), 127.82 (CH ꢃ 3), 127.77 (CH ꢃ 3), 127.7 (CH), 127.5
(CH ꢃ 3), 127.4 (CH ꢃ 3), 127.3 (CH ꢃ 3), 127.2 (CH), 123.0
(CH), 112.4 (CH), 98.8 (CH), 79.0 (CH), 76.5 (CH), 74.73 (CH),
74.70 (CH2), 73.4 (CH2), 73.0 (CH2), 72.9 (CH2), 72.3 (CH), 70.8
(CH2), 70.3 (CH2), 69.4 (CH), 68.8 (CH2), 68.0 (CH2), 52.3 (CH),
36.7 (CH2), 31.9 (CH2 ꢃ 3), 29.7 (CH2 ꢃ 20), 29.50 (CH2), 29.45
(CH2), 29.4 (CH2), 29.3 (CH2 ꢃ 2), 29.2 (CH2), 25.6 (CH2), 22.7
(CH2 ꢃ 3), 14.1 (CH3 ꢃ 2); HRMS (ESI, M + H+) calcd for
Acknowledgements
The authors thank the Ministry of Science and Technology
(MOST) in Taiwan (NSC101-2113-M-005-006-MY2) and National
Chung Hsing University for nancial support. The authors
special thank Professor Dr Alice L. Yu and post-doctoral fellow
Dr Jung-Tung Hung of Academia Sinica, Taipei 115, Taiwan, for
performing the bioassay of analogue 2.
C
92H131O9NNa 1416.9716, found 1416.9722.
(2S,3S,5S)-2-Hexacosanoylamino-1-O-(a-d-galactopyranosyl)-
Notes and references
octadec-1,3,5-triol (2). Compound 13 (118 mg, 0.08 mmol) was
dissolved in MeOH/CHCl3 (3/1 ratio, 2 mL) at room tempera-
ture. Pd(OH)2/C (118 mg, Degussa type) was added to the reac-
tion mixture, the reaction vessel was purged with hydrogen, and
the mixture was stirred under 60 psi pressure at the same
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26534 | RSC Adv., 2014, 4, 26524–26534
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