Synthesis of serine-based glycolipids as potential TLR4 activators

Article abstract of DOI:10.1039/c0ob00990c

A new series of monosaccharide-based glycolipids devoid of phosphate groups and with two lipid chains were rationally designed by varying the lipid chain lengths and saccharide structure of a α-GalCer-derived lead compound (CCL-34) that is a potent TLR4 agonist. The NF-κB activity of a 60-membered galactosyl serine-based synthetic library containing compounds with various lipid chain lengths was measured in a HEK293 cell line that stably expressed human TLR4, MD2, and CD14 (293-hTLR4/MD2-CD14). The results showed that the optimal carbon chain lengths for the lipid amine and fatty acid to activate TLR4 were 10-11 and 12, respectively. Evaluation of a 20-membered synthetic glycosyl serine-based lipid library containing compounds with various saccharide moieties and fixed lipid chain lengths revealed that the galactose moiety in CCL-34 could be replaced by glucose without loss of activity (CCL-34-S3 and CCL-34-S16). Changing the orientation of the anomeric glycosidic bond of CCL-34 resulted in a complete loss of activity (β-CCL34). Surprisingly, a change in configuration of the anomeric glycosidic bond in a glucosyl glycolipid is tolerable (CCL-34-S14). Another noteworthy observation is that the activity of a l-fucosyl derived glycolipid (CCL-34-S13) was comparable to that of CCL-34. In sum, this study determines the structural features that are crucial for an optimal TLR4-stimulating activity. It also provides several molecules with immunostimulating potential.

Full text of DOI:10.1039/c0ob00990c

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PAPER
Synthesis of serine-based glycolipids as potential TLR4 activators†
Li-De Huang,a Hong-Jyune Lin,a Po-Hsiung Huang,b Wei-Chen Hsiao,a L. Vijaya Raghava Reddy,a
Shu-Ling Fu*b and Chun-Cheng Lin*a
Received 6th November 2010, Accepted 22nd December 2010
DOI: 10.1039/c0ob00990c
A new series of monosaccharide-based glycolipids devoid of phosphate groups and with two lipid
chains were rationally designed by varying the lipid chain lengths and saccharide structure of a
a-GalCer-derived lead compound (CCL-34) that is a potent TLR4 agonist. The NF-kB activity of a
60-membered galactosyl serine-based synthetic library containing compounds with various lipid chain
lengths was measured in a HEK293 cell line that stably expressed human TLR4, MD2, and CD14
(293-hTLR4/MD2-CD14). The results showed that the optimal carbon chain lengths for the lipid
amine and fatty acid to activate TLR4 were 10–11 and 12, respectively. Evaluation of a 20-membered
synthetic glycosyl serine-based lipid library containing compounds with various saccharide moieties
and fixed lipid chain lengths revealed that the galactose moiety in CCL-34 could be replaced by glucose
without loss of activity (CCL-34-S3 and CCL-34-S16). Changing the orientation of the anomeric
glycosidic bond of CCL-34 resulted in a complete loss of activity (b-CCL34). Surprisingly, a change in
configuration of the anomeric glycosidic bond in a glucosyl glycolipid is tolerable (CCL-34-S14).
Another noteworthy observation is that the activity of a L-fucosyl derived glycolipid (CCL-34-S13) was
comparable to that of CCL-34. In sum, this study determines the structural features that are crucial for
an optimal TLR4-stimulating activity. It also provides several molecules with immunostimulating
potential.
4 (TLR4), a key member of pattern recognition receptors that
are involved in the induction of innate and adaptive immune
Introduction
Glycolipids1 are components of the ubiquitous membrane that
is found in many species. In general, they are constituted by a
hydrophobic lipid group and a hydrophilic carbohydrate moiety,
which is linked to the terminal hydroxyl group of sphingosine
or glycerol/inositol. Glycolipids are essential components of cell
membranes in the body; they are primarily located in the outer
layer of the plasma membranes, where they interact with the
extracellular environment. Thus, glycolipids play important roles
in the regulation of cell-growth and development. They also
serve as important epitopes in cell recognition, e.g., in cell-cell
communication2 and microbial infections.3,4 In addition, recent
studies have shown that glycolipids may serve as immunomodula-
tors with therapeutic potential for Parkinson’s disease, Alzheimer’s
disease, psoriasis, AIDS treatment, and antiviral infections.5–7
Lipopolysaccharide (LPS),8 an amphiphilic glycolipid located
in the outer membrane of Gram-negative bacteria, is associated
with pathogen recognition by the host innate immune system.
Cellular recognition for LPS is mediated by Toll-like receptor
responses.9 Structurally, LPS is composed of a polysaccharide and
lipid A; the latter is involved in many LPS-mediated responses.10
Indeed, lipid A is the main region of LPS that interacts with
TLR4 and can trigger innate immune systems.11 It is thought
that TLR4 agonists are potent immunostimulators with multiple
therapeutic applications, such as vaccine adjuvants and cancer
immunotherapeutic agents. On the other hand, TLR4 antagonists
may be therapeutically useful for inflammatory diseases.12 Sub-
stantial efforts have been devoted to the synthesis of molecules that
mimic lipid A and can serve as agonists or antagonists for TLR4
activation processes.13,14 Most of the currently available bioactive
lipid A analogs contain a phosphate saccharide scaffold with lipid
substituents.15–18 Such molecules (e.g., monophosphoryl lipid A
(MPL)19 and aminoalkylglucosaminide 4-phosphates (AGPs)20)
have yielded promising results in clinical trials in which they were
used as therapeutic vaccines/adjuvants against cancer, infectious
diseases, and allergies. Interestingly, it was recently found that
some compounds with structures totally different to that of lipid
A, e.g., taxol,21 flavolipin,22 vitamin D,23 and lipids linked to a
phosphate-containing acyclic backbone,24–26 exhibited activities
towards TLR4 that were similar to that of LPS.
aDepartment of Chemistry, National Tsing Hua University, 101 Sec. 2, Kuang
Fu Rd, Hsinchu 30013, Taiwan
bInstitute of Traditional Medicine, National Yang-Ming University, 155
Sec.2, Li-Nong St., Taipei 11221, Taiwan
In our continuous efforts27–29 to develop a a-GalCer analog
as a vaccine adjuvant, we have found that serine-based a-
GalCer analogs could activate the TLR4 signaling pathway.
† Electronic supplementary information (ESI) available: Bioactivities and
NMR spectra of synthesized compounds. See DOI: 10.1039/c0ob00990c
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In our previous studies,30 among the forty-seven serine-based
glycolipids tested, two a-GalCer analogs (CCL-34 and CCL-
25) stimulated a TLR4-dependent NF-kB activity in a murine
macrophage cell line RAW 264.7 and in a human HEK293
cell line that stably expressed TLR4, MD2, and CD14 (293-
hTLR4/MD2-CD14). No activity was detected for a-GalCer.
Furthermore, TNF-a production in CCL-34-treated primary
bone marrow cells was also found to be TLR4-dependent. These
results are interesting for the development of a new class of potent
monosaccharide immunomodulators.30 As the structure of CCL-
34 is relatively simple and the synthesis of serine-based glycolipids
is straightforward, CCL-34 can be easily derivatized to obtain
a new generation of compounds. To study the structure–activity
relationship of serine-based glycolipids, we have synthesized many
serine-based derivatives of CCL-34 by either changing the lipid or
the carbohydrate moiety. The bioactivity results showed that the
optimal chain length for the lipid moiety was eleven or twelve
carbons and that the galactose in CCL-34 could be replaced by
glucose or fucose without losing any activity.
separated, the para-methoxybenzyl protecting groups of 3 were
replaced by acetyl groups to give the separable compounds 4
and 5. Deprotection of the allyl group of 4 using Pd(PPh3)4
followed by amidation with five different lipid amines containing
ten- to fourteen-carbon linear chains afforded compounds 4a–4e
in 71–84% yields. Removal of the Fmoc group with piperidine
in CH2Cl2, coupling with different fatty acids, and deacetylation
gave sixty serine-based galactosyl lipids with total yields of 54–65%
over three steps. The NF-kB activation assays were performed in
293-hTLR4/MD2-CD14 cells. The results revealed that the most
effective lengths of fatty acid chain and lipid amine for NF-kB
activation were eleven and twelve carbons, respectively (CCL-34;
named as 4b-3 in Scheme 1 and S1 in supporting information).
Although the compound 4a-3 containing a ten-carbon lipid amine
showed a NF-kB-stimulating activity comparable to that of 4b-
3, compounds with other chain lengths had significantly lower
activities (see supporting information S3).
Development of a-selective galactosylation
Even though galactosyl lipids could be obtained following the
synthetic procedures described in Scheme 1, the moderate a-
selectivity of galactosylation and the difficulty in separating the
a/b mixture hampered further investigations on the structure–
activity relationship of the carbohydrate moiety on the glycolipid
activity. We therefore developed a new glycosylation strategy that
led to a higher a-selectivity. As previously reported, an exclusive a-
selectivity (Table 1, entry 1) can be achieved by using a phosphite 6
as the glycosyl donor.31 However, the preparation of the saccharide
donor itself is tedious and time-consuming, as it requires nine steps
(from galactose). The use of thiocresol as the leaving group in
the donor 7 with PMB (p-methoxybenzyl) as hydroxyl protecting
group did not lead to an acceptable a-selectivity with Fmoc-Ser-
OAll as the glycosylation acceptor (Table 1, entries 2 and 3).
However, when compound 8 was used as the donor, the desired a-
glycoside was obtained in 55% yield at low temperature (-40 ◦C to
RT) and in 63% yield at room temperature (Table 1, entries 4 and
5). To further improve the yield of the a-anomer, Gervay-Hague’s
method22 was adopted because it has been shown to give a high
yield and a high a-selectivity in galactosylation reactions. Reaction
between the donor 9 and Fmoc-Ser-OAll as the acceptor led to
the degradation of the starting material during the generation of
iodide in the first step (Table 1, entry 6). This may be due to
the undesired deprotection of the acid-sensitive p-methoxybenzyl
group by iodotrimethylsilane (TMSI). The reaction between the
iodide donor generated in situ from galactosyl acetate 1032 and
TBAI and Fmoc-Ser-OAll (1.2 equiv) acceptor in the presence of
Results and discussion
Synthesis of serine-based galactosyl lipids
We had previously synthesized a library of 47 serine-based galac-
tosyl lipids30 and had found that the length of both the fatty acid
and the lipid amine that is linked to serine had a profound effect
on the activation of TLR4. To determine the structure–activity
relationship of serine-based galactosyl lipids, a new synthetic
library featuring lipids of different lengths and various sugar
moieties was synthesized. As shown in Scheme 1, the synthesis
of compound 4 started with the glycosylation of the donor 2 with
the Fmoc-Ser-OAll acceptor to give a galactosyl serine mixture
3 (85% yield, a/b = 2.5/1). As the a/b anomers could not be
˚
DIPEA, 4 A MS, and toluene gave the galactoside 10a in 60% yield
with an exclusive a-selectivity (Table 1, entry 7). When the same
method was applied to glucose, 3- and 4-allylated galactose, and
fucose (Table 1, entries 8–11), the a-anomers were obtained with
high selectivity. The good a-selective galactosylation procedure
allowed us to modify the carbohydrate moiety of galactosyl lipids
for further SAR investigations.
˚
Scheme 1 Reagents and conditions: (a) DDP, 1-H-tetrazole, DCM, 4 A
MS, rt; (b) Fmoc-Ser-OAll, TMSOTf, DCM, 4 A MS, 0 ◦C, 85% for
˚
Initially, we focused on the modification of the galactose moiety
of CCL-34. For example, we replaced the C6 hydroxyl group with
an amino or carboxylic group. We also O-alkylated the C3 or
C4 hydroxyl group and changed the sugar moiety to glucose.
As shown in Scheme 2, deprotection of the allyl group of 10a,
two steps; (c) TFA, DCM; (d) Ac2O, Pyr., DMAP, 70% for two steps; (e)
Pd(PPh3)4, MeNHPh, THF; (f) lipid amine, EDC, HOBt, NMM, DCM,
71–84% for two steps; (g) piperidine, DCM; (h) fatty acid, EDC, HOBt,
TEA, DCM, 65–73%; (i) NaOMe, MeOH, 85–90%. The structures of
compounds 4a-1 to 4e-12 are shown in supporting information S1.
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Table 1 Studies of a-selective galactosylation
oxidizing the primary hydroxyl group in CCL-34 or deprotecting
the benzyl groups prior to the esterification of the acid, resulted
in relatively low yields and purification difficulties. Glucosyl lipids
CCL-34-S3, CCL-34-S4, and CCL-34-S5 were obtained in good
yields following a similar synthetic procedures described above.
In addition to the oxidation of the C6 hydroxyl group of CCL-
34, amination of the C6 position of galactose was also performed
(Scheme 3). The direct transformation of the C6 hydroxyl group of
15 into an amino group resulted in an incomplete reaction, which
may be due to the steric hindrance of the C4 benzyl protection. In
order to circumvent the above problem, compound 18 with a free
C4 hydroxyl group was synthesized starting from 8a. Selective
deprotection of the 4,6-O-benzylidene group in 17, which was
synthesized in 53% yield (over four steps) from 8a, was achieved
by using VO(OTf)233 and gave compound 18. Tosylation of the C6
hydroxyl group of 18 followed by a SN2 reaction with sodium azide
and the reduction of the azido group gave the amine 19.
entry
Condition
donor
% yield
Product (a/b)
1
2
3
4
5
6
7
8
A
B1
B2
B3
B4
C
C
C
C
C
6
95
71
95
55
63
6a (a only)
3a/b (2 : 1)
7a/b (1.5 : 1)
8a (a only)
8a (a only)
—
10a (a only)
11a (a only)
12a (a only)
13a (a only)
14a (a only)
7
7
8
8
a
8
—
10
11
12
13
14
60
54
66
57
56
9
10
11
C
a Starting material was decom◦posed during reaction. Condition A: TfOH,
˚
˚
CH2Cl2, 4 A MS, 60 min, 0 C–RT; B1: NIS, TfOH, CH2Cl2, 4 A MS,
10 min, RT; B2: NIS, TfOH, CH2Cl2, 4 A MS, 120 min, -40 ◦C; B3: NIS,
˚
◦
˚
˚
TfOH, CH2Cl2, 4 A MS, 60 min, -40 C–RT; B4: NIS, TfOH, CH2Cl2, 4 A
MS, 120 min, RT; C: (i) TMSI, 4 A MS, CH2Cl2, 10 min, 0 ◦C; (ii) TBAI,
˚
DIPEA, toluene, 4 A MS, 180 min, 80 ◦C.
˚
Scheme 3 Reagents and conditions: (a) Pd(PPh3)4, MeNHPh, THF; (b)
n-undecylamine, EDC, HOBt, NMM, DCM; (c) piperidine, DCM; (d)
lauric acid, EDC, HOBt, TEA, DCM, 53% for four steps; (e) VO(OTf)2,
MeOH–MeCN–DCM (2 : 6 : 1), 76%; (f) TsCl, Pyridine; (g) NaN3, DMF,
71% for two steps; (h) Pd/C, H2, DCM–MeOH, 86%; (i) TFA, DCM; (j)
R3-acid, EDC, HOBt, TEA, DCM.
Acylation of the amino group in the presence of EDCI, HOBt,
and TEA in CH2Cl2 with different acids and deprotection of the
PMB groups with TFA in CH2Cl2 afforded CCL-34-S7, CCL-34-
S8, CCL-34-S9, and CCL-34-S10 in good yields. Deprotection
of the PMB groups in 19 with TFA provided CCL-34-S6 in 80%
yield.
The alkylation of the C4 and C3 hydroxyl groups of the galactose
moiety of CCL-34 was then undertaken (Scheme 4). The desired
lipid chains were introduced as described in Scheme 1. Reduction
of the benzyl groups and unsaturation of the C4 allyl group using
H2 and Pd/C in MeOH followed by deprotection of the PMB and
benzylidene groups gave CCL-34-S11 and CCL-34-S12 in 43%
and 34% (over 5 steps), respectively. Similarly, the fucosyl lipid
CCL-34-S13 was obtained from 14a in five steps with 57% overall
yield.
Scheme 2 Reagents and conditions: (a) Pd(PPh3)4, MeNHPh, THF;
(b) n-undecylamine, EDC, HOBt, NMM, DCM; (c) piperidine, DCM;
(d) lauric acid, EDC, HOBt, TEA, DCM; (e) NaOMe, MeOH, DCM;
(f) TEMPO, BAIB, DCM-H2O (2 : 1); (g) MeI, Cs2CO3, DMF; (h) H2,
Pd/C, MeOH, DCM; (i) LiOH, H2O–THF–MeOH (1 : 2 : 1); (j) Pd/C,
H2, MeOH, DCM.
coupling with n-undecylamine, deprotection of the Fmoc group,
coupling with lauric acid, and deacetylation gave compound 15
in an overall yield of 68%. A TEMPO-mediated oxidation of 15
generated the corresponding sugar acid that was esterified using
MeI and Cs2CO3 in DMF and then full deprotected to give CCL-
34-S1 in 76% yield (over three steps). De-esterification of CCL-
34-S1 under mild reaction conditions in the presence of LiOH
gave the sugar acid CCL34-S2 in 83% yield. It is noteworthy that
both methods tested to obtain CCL34-S2, i.e., either selectively
The orientation of the glycosidic linkage might influence the
interaction between the molecule and the protein receptor. To
study the importance of the a-orientation of the glycosidic bond
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Scheme 4 Reagents and conditions: (a) piperidine, DCM; (b) lauric
acid, EDC, HOBt, TEA, DCM; (c) Pd(PPh3)4, MeNHPh, THF; (d)
n-undecylamine, EDC, HOBt, TEA, DCM; (e) H2, Pd/C, MeOH, DCM;
(f) TFA, DCM.
in serine-based glycolipids, the b-galactosyl lipid b-CCL-34 and
the b-glucosyl lipid CCL-34-S14 were synthesized from 5 and 20
in 43% and 55% overall yields, respectively (Scheme 5).
Scheme 7 Reagents and conditions: (a) HBr, HOAc; (b) Fmoc-Ser-OAll,
˚
AgOTf, DCM, 4 A MS, 86%; (c) Pd(PPh3)4, MeNHPh, THF; (d)
undecylamine, EDC, HOBt, TEA, DCM, 80% for two steps; (e) piperidine,
DCM, 90%; (f) lauric acid, EDC, HOBt, TEA, DCM, 80%; (g) Zn, HOAc,
THF, 95%; (h) NaOMe, MeOH, DCM, 60%; (i) lauric acid, EDC, HOBt,
TEA, DCM, 54%; (j) NaOMe, MeOH, DCM, 60%; (k) Zn, Ac2O, HOAc,
THF, 72%; (l) NaOMe, MeOH, DCM, 82%.
Scheme 5 Reagents and conditions: (a) Pd(PPh3)4, MeNHPh, THF; (b)
n-undecylamine, EDC, HOBt, NMM, DCM; (c) piperidine, DCM; (d)
lauric acid, EDC, HOBt, TEA, DCM; (e) NaOMe, MeOH.
To investigate whether the alteration of the sugar moiety affects
the activity of CCL-34, the galactose of CCL-34 was replaced with
different monosaccharides, such as mannose and glucosamine. As
shown in Scheme 6, glycosylation of the peracetylated mannoside
21 with Fmoc-Ser-OH using BF3·OEt2 as the promoter gave the a-
anomer 22 in 67% yield. Compound 22 was then transformed into
CCL-34-S15 by following the standard five-step sequence (34%
overall yield).
THF gave the free amine 26 (95% yield). This amine was then
deacetylated in the presence of NaOMe and MeOH to give CCL-
34-S16 in 60% yield. Coupling of the free amino group in 26 with
lauric acid followed by deacetylation afforded CCL-34-S17 with
three lipid chains in 32% yield. On the other hand, deprotection
of the Troc group in the presence of acetic anhydride gave the N-
acetyl protected product 27 in 72% yield. Removal of the O-acetyl
groups in 27 afforded the N-acetylated glycolipid CCL-34-S18 in
59% yield. Finally, in order to determine the importance of the
carbohydrate part for the immunostimulating activity of CCL-34,
a sugarless serine lipid CCL-34-S19 (shown in Scheme 8) was also
synthesized using the procedures described in Scheme 1.
Scheme 6 Reagents and conditions: (a) Fmoc-Ser-OH, BF3–OEt2, DCM,
67%; (b) EDC, HOBt, n-undecylamine, NMM, DCM, 87%; (c) piperidine,
DCM; (d) lauric acid, EDC, HOBt, TEA, DCM, 73%, two steps; (e)
NaOMe, MeOH, 80%.
A literature survey of TLR-4 ligands, lipid A, and its mimics14,34
reveals that the polar head group is mostly GlcNAc and that
the lipid moiety is linked to the C2 or C3 position of GlcNAc
This implies that a lipid extended from the C2 position of a
glycolipid sugar may interact with a receptor and affect the
immune response. Thus, GlcNAc serine-based glycolipids were
also synthesized (Scheme 7). The anomeric acetate of the known
compound 2335 was converted into bromide and then coupled
with Fmoc-Ser-OAll to yield 24 (86% yield over two steps). The
precursor 25 for the synthesis of compounds CCL-34-S16, S17,
and S18 was obtained in 57% yield by using the linear four-step
sequence previously described. Selective deprotection of the Troc
group at position C2 in compound 25 using Zn and HOAc in
Scheme 8 Reagents and conditions: (a) n-undecylamine, EDC, HOBt,
DIPEA, DCM, 93%; (b) piperidine, DCM; (c) lauric acid, EDC, HOBt,
DIPEA, DCM, 71% for two steps.
Bioactivities of glycosyl serine-based lipids: SAR analysis
The strategy to identify TLR4 activators has been described in
detail in our previous study.30 In short, 293-hTLR4/MD2-CD14
cells seeded in 24-well plates at a density of 2¥ 105 cells per well
were grown overnight and treated for 5 h with LPS (10 ng ml-1; the
positive control) or the synthesized glycosyl lipids (10 mM). The
glycosyl lipids that were able to activate NF-kB, a downstream
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signaling molecule of the TLR4 pathway, were identified using a
NF-kB activity-based reporter assay as described previously.
Fig. 1 summarizes the NF-kB activity induced by CCL-34 and
the synthesized glycosyl lipid derivatives. The lack of activity
of the serine lipid CCL-34-S19 suggests that the sugar part
is indispensable for TLR4 stimulation. Of all the synthesized
glycosyl lipids tested, five derivatives (i.e., CCL-34-S2, CCL-
34-S3, CCL-34-S13, CCL-34-S14,and CCL-34-S16) displayed
significant NF-kB activities. The glucosyl lipid derivative CCL-
34-S3 induced a NF-kB activation that was similar to that of
CCL-34, suggesting that the configuration of the 4-hydroxyl group
is not crucial for activity. Although the b-anomer of CCL-34
did not show any activity, the glucosyl lipid derivative with a
b-anomeric configuration (CCL-34-S14) still induced significant
NF-kB activation. The glucosamine lipid derivative CCL-34-S16
showed a good activity, but any further derivatization of the free
amino group led to a complete loss of NF-kB activity, as clearly
evidenced in the cases of CCL-34-S17 and CCL-34-S18. On the
other hand, the mannosyl lipid derivative CCL-34-S15 with a
change in configuration of the C2 hydroxyl group showed 60%
of the NF-kB activity of CCL-34, indicating the importance of
the equatorial configuration of the C2 functional group on the
sugar moiety. It is noteworthy that the L-fucosyl lipid derivative
CCL-34-S13 has a NF-kB activity comparable to that of CCL-34.
Nevertheless, the C3-O-alkylated derivatives of CCL-34, namely
CCL-34-S12, did not give promising results.
glycolipid is inserted and (2) the decrease of the strength of
the hydrogen bonding or dipole–dipole interaction between the
hydroxyl groups of the saccharide moiety and the receptor when
the hydroxyl groups were transformed into an ethoxy or amino
group. The modification of the C4-hydroxyl group resulted in
promising results and further investigation is needed.
Conclusions
In conclusion, we have synthesized a library of serine-based lipids
with high efficiency and demonstrated that an acid (Rx) with 12
carbons and an amine (Ry) with 11 carbons are the most effective
components for TLR4 activation. In addition, the synthesis of
19 serine-based glycolipids with different saccharide moieties and
orientations of glycosidic linkage was also achieved. The bioassay
results revealed that out of the 19 compounds synthesized, five
compounds possessed an activity that was comparable to that
of CCL-34. It is noteworthy that the serine-based b-glucosyl lipid
CCL-34-S14 could be easily synthesized in six steps with an overall
yield of 30%. This compounds displayed a TLR4-stimulating
activity that was comparable to that of CCL-34, suggesting that
this glycolipid is a potential immunostimulator, although further
studies are required.
Experimental
Compounds 1,36 2, 3a,30 3b, 4, 5,37 6,31 6a,31 7,36 8,38 10,39 11,40 14,41
20,37 22,42 and 2335 were previously reported. Our prepared samples
1
showed consistent H and 13C NMR spectral data. All reagents
were purchased from commercial suppliers and used without
further purification unless otherwise noted. Dichloromethane
(DCM) and toluene were distilled from calcium hydride under
N2. Tetrahydrofuran (THF) was distilled under N2 from sodium
˚
prior to use. Powdered molecular sieves (4 A MS) were activated
in vacuo at 150 ◦C for 8 h and cooled to room temperature
in vacuo prior to use. Analytical thin layer chromatography was
performed on E. Merck silica gel 60 F254 plates (0.25 mm).
Liquid column chromatography was performed using a forced
flow of the indicated solvent on silica gel 60 (E. Merck Co.).
NMR spectra were recorded on Bruker 400 Hz instruments and
are reported in parts per million (d) units. Coupling constants
(J) are reported in hertz (Hz). 1H NMR spectra were recorded in
CDCl3 and referenced to residual CHCl3 at 7.24 ppm. 13C NMR
spectra were referenced to the central peak of CDCl3 at 77.0 ppm.
Assignment of 1H NMR signals was achieved using 2D methods
(COSY). High-resolution mass spectra were obtained by means
of a Micromass (Autospec) mass spectrometer. All reactions were
carried out in oven-dried glassware (104 ◦C) under an atmosphere
of nitrogen unless otherwise indicated.
Fig.
1 The activities of glycolipids on NF-kB activation in
293-hTLR4/MD2-CD14 cells.
The biological results also indicate that the galacturonic acid
glycolipid CCL-34-S2 has an activity similar to that of CCL-
34. However, its methyl ester derivative CCL-34-S1 did not have
any activity. Substitution of the C6 hydroxy group by an amino
group (CCL-34-S6) and amidation (CCL-34-S7–S10) resulted in
a significant decrease in activity, indicating that the modification
of the C6 hydroxyl group of galactose was acceptable but that
any elongation at this location was detrimental. In contrast,
the glucuronic acid glycolipid CCL-34-S5 and its methyl ester
derivative CCL-34-S4 did not have any activity, indicating that
the C6 hydroxyl group cannot be modified on a glucose lipid
derivative.
N-(9-Fluorenylmethoxycarbonyl)-O-(2,3-di-O-para-methoxy-
benzyl-4,6-O-benzylidene-a-D-galactopyranosyl)-L-serine allyl
ester (8a)
Based on the above-described structure–activity-relationship
studies, it can be concluded that the C2, C3 and C6 positions of
the saccharide moiety cannot be further modified to obtain new
derivatives with improved activity. The observed loss of activity
may be attributed to (1) crowding in the pocket in which the
A mixture of the glycosyl donor 8 (2.15 g, 3.49 mmol), Fmoc-Ser-
˚
OAll (1.15 g, 3.13 mmol), and 4 A MS (2 g) in dried CH2Cl2 (30 mL)
was stirred for 10 min at room temperature under N2. A solution
of NIS (1.46 g, 6.49 mol) in dried CH2Cl2 (5 mL) and diluted
with TfOH (85 mL, 0.97 mmol) was then added sequentially. The
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reaction mixture was stirred at room temperature for 0.5 h and
monitored by TLC. After the reaction was complete (based on
TLC analysis), the molecular sieves were removed by filtration
over Celite. The filtrate was diluted with EtOAc and the organic
layer was washed with saturated aqueous Na2S2O3, NaHCO3, and
brine. The organic layer was dried over MgSO4, concentrated, and
subjected to a flash silica gel column chromatography to give 8a
(exclusive a-anomer) as a white foam (1.69 g, 1.97 mmol, 63%). Rf
0.14 (1 : 2 EtOAc–hexane); 1H (400 MHz, CDCl3) d 3.63 (1H, br
s), 3.76 (3H, s), 3.77 (3H, s), 3.82 (1H, dd, J = 3.0, 11.3 Hz), 3.87
(1H, dd, J = 3.0, 10.1 Hz), 3.90 (1H, d, J = 13.4 Hz), 4.03 (1H,
dd, J = 3.4, 10.1 Hz), 4.13 (1H, d, J = 3.0 Hz), 4.16–4.23 (3H, m),
4.34 (1H, dd, J = 7.2, 10.6 Hz), 4.41 (1H, dd, J = 7.2, 10.6 Hz),
4.48–4.52 (1H, m), 4.54 (1H, d, J = 10.2), 4.60 (1H, dd, J = 10.1,
3.0 Hz), 4.65 (1H, d, J = 11.3 Hz), 4.72 (1H, d, J = 10.2 Hz), 4.74
(1H, d, J = 11.3 Hz), 4.80 (1H, d, J = 3.4 Hz), 5.20 (1H, d, J = 10.5
Hz), 5.30 (1H, d, J = 17.2 Hz), 5.45 (1H, s), 5.81–5.91 (1H, m), 6.16
(1H, d, J = 8.4 Hz), 6.84 (4H, dd, J = 6.5, 8.5 Hz), 7.25–7.40 (11H,
m), 7.50 (1H, d, J = 1.3 Hz), 7.52 (1H, d, J = 1.9 Hz), 7.58 (2H,
d, J = 7.4 Hz), 7.75 (2H, d, J = 7.6 Hz); 13C (100 MHz, CDCl3) d
46.90, 54.62, 55.04, 63.19, 66.05, 67.04, 69.08, 70.31, 71.51, 73.11,
74.32,74.82, 75.08, 100.18, 100.74, 113.56, 113.60, 118.55, 119.86,
124.87, 124.95, 126.14, 126.92, 126.96, 127.57, 127.93, 128.69,
129.10, 129.37, 130.49, 130.54, 131.36, 137.66, 141.09, 143.58,
155.92, 159.09, 169.76; HRMS (ESI) calc. for C50H50NO12 [M -
H]+ 856.3333, found 856.3334.
126.72, 127.15, 127.31, 127.36, 127.47, 128.04, 128.09, 131.22,
137.83, 138.08, 138.24, 140.93, 143.49, 155.63, 169.30, 170.23;
HRMS (ESI) calc. for C50H52NO11 [M + H]+ 842.3540, found
842.3522.
General procedure for the deprotection of the allyl ester and the
amidation
N-Methylaniline (3 mmol) was added to a solution of serine
glycoside (1 mmol) and Pd(PPh3)4 (0.3 mmol) in THF (10 mL).
The reaction mixture was stirred at room temperature for 2 h,
and then the solvent was removed. The crude was diluted with
EtOAc, washed by water, 10% aqueous HCl, brine, dried with
MgSO4, concentrated, and subjected to a short pad of silica gel to
remove residual N-methylaniline. The crude product was dissolved
in dry CH2Cl2 (10 mL) and EDCI (2 mmol), HOBt (2 mmol),
lipid amine (1.5 mmol), and NMM (2 mmol) were successively
added. The reaction mixture was stirred at room temperature
for 2 h. The solvent was removed in vacuo and the residue was
diluted with EtOAc and washed with 10% aqueous HCl, saturated
aqueous NaHCO3, and brine. The organic layer was dried over
MgSO4, concentrated, and subjected to a flash silica gel column
chromatography to give the desired product.
General procedure for the deprotection of the Fmoc group and the
amidation
Piperidine (1 mL) was added to a solution of the Fmoc-protected
glycoside (1 mmol) in CH2Cl2 (10 mL). The reaction mixture was
stirred at room temperature for 20 min and then the solvent was
removed. The crude was subjected to silica gel column chromatog-
raphy to give the deprotected product that was dissolved in dry
CH2Cl2 (10 mL). EDCI (2 mmol), HOBt (2 mmol), lipid acid (1.5
mmol), and DIPEA (2 mmol) were successively added and the
reaction mixture was stirred at room temperature for 3 h. The
reaction mixture was then diluted with CH2Cl2 and washed with
10% aqueous HCl, saturated aqueous NaHCO3, and brine. The
organic layer was dried over MgSO4, concentrated, and subjected
to a flash silica gel column chromatography to give the di-lipid
product.
N-(9-Fluorenylmethoxycarbonyl)-O-(3,4,5-tri-O-benzyl-6-
acetoxy-a-D-galactopyranosyl)-L-serine allyl ester (10a)
TMSI (0.3 mL, 2.10 mmol) was added to a solution of the glycosyl
˚
acetate (1.01 g, 1.89 mmol) and 4 A MS (1 g) in CH2Cl2 (20 mL) at
0 ◦C. The reaction mixture was stirred at 0 ◦C for 20 min and then
3 mL of anhydrous toluene were added. The molecular sieves were
removed by filtration over Celite. After concentrating the filtrate
and azeotroping three times with toluene, the obtained yellow
residue was dissolved in toluene (3 mL) and kept under N2. In
˚
a separate flask, 4 A MS, TBAI (1.42 g, 3.84 mmol), Fmoc-Ser-
OAll (0.50 g, 1.37 mmol), and DIPEA (0.50 mL, 3.03 mmol) were
mixed in toluene (20 mL). The mixture was stirred under N2 at
◦
80 C for 20 min. Glycosyl iodide was then cannulated into the
O-(2,3,4-Tri-O-benzyl-6-hydroxy-a-D-galactopyranosyl)-N-
reaction mixture. After the reaction mixture was stirred at 80 ◦C
for 3 h. The reaction was stopped by adding EtOAc (10 mL) and
cooling to 0 ◦C. The white precipitate and the molecular sieves
were removed by filtration over Celite. The filtrate was washed
with saturated aqueous Na2S2O3 and brine. The organic layer was
dried over MgSO4, concentrated, and subjected to a flash silica
gel column chromatography to give 10a (exclusive a-anomer) as a
white foam (0.69 g, 0.82 mmol, 60%). Rf 0.38 (1 : 1 EtOAc–hexane);
1H (400 MHz, CDCl3) d 1.94 (3H, s), 3.83 (1H, dd, J = 2.9, 11.2
Hz), 3.86–3.90 (3H, m), 4.05 (1H, dd, J = 3.5, 10.6), 4.09–4.18 (3H,
m), 4.21 (1H, t, J = 7.0 Hz), 4.38 (2H, d, J = 7.0 Hz), 4.53–4.56
(1H, m), 4.58–4.64 (4H, m), 4.73–4.78 (3H, m), 4.85 (1H, d, J =
11.7 Hz), 4.97 (1H, d, J = 11.4 Hz), 5.18 (1H, dd, J = 1.1, 10.3 Hz),
5.28 (1H, dd, J = 1.1, 17.2 Hz), 5.85 (1H, ddd, J = 5.7, 10.3, 17.2
Hz), 6.14 (1H, d, J = 8.5 Hz), 7.25–7.40 (19H, m), 7.59 (2H, d, J =
7.5 Hz), 7.73 (2H, d, J = 7.6 Hz); 13C (100 MHz, CDCl3) d 20.37,
46.76, 54.43, 63.31, 65.76, 66.70, 69.06, 70.04, 72.94, 74.24, 74.38,
76.01, 76.68, 77.00, 77.32, 78.18, 99.14, 118.30, 119.64, 124.76,
dodecanoyl-L-serine undecylamide (15)
Compound 15 was obtained in 68% from 10a as a white foam.
Rf 0.25 (1 : 1 EtOAc–hexane); 1H (400 MHz, CDCl3) d 0.86 (6H,
t, J = 6.6), 1.21–1.27 (34H, m), 1.50–1.66 (2H, m), 2.16 (2H, t,
J = 7.9), 2.63–2.71 (1H, m), 3.04–3.12 (1H, m), 3.43 (1H, dd, J =
9.1, 11.5), 3.47 (1H, dd, J = 4.8, 11.1), 3.72 (1H, dd, J = 7.1,
11.1), 3.79–3.88 (3H, m), 3.93 (1H, dd, J = 4.4, 11.5), 4.11 (1H,
dd, J = 3.7, 10.0), 4.39 (1H, ddd, J = 4.4, 6.6, 9.1), 4.61 (1H,
d, J = 11.6), 4.69 (1H, d, J = 10.8), 4.75 (1H, d, J = 11.8), 4.85
(1H, d, J = 11.8), 4.85 (1H, d, J = 10.8), 4.93 (1H, d, J = 11.6),
5.12 (1H, d, J = 3.7), 6.66 (1H, d, J = 6.6), 6.94 (1H, t, J = 5.4),
7.26–7.40 (15H, m); 13C (100 MHz, CDCl3) d 13.94, 22.50,25.37,
26.72, 29.16, 29.34, 29.44, 31.73, 36.25, 39.39, 51.48, 62.05, 68.48,
71.46, 72.81, 74.41, 76.41, 79.34, 98.37, 127.18, 127.55, 127.67,
127.96, 128.14, 128.21, 128.30, 128.35, 137.46, 138.08, 138.21,
169.42, 173.15; HRMS (ESI) calc. for C53H80N2O8Na [M + Na]+
895.5812, found 895.5818.
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N-(9-Fluorenylmethoxycarbonyl)-O-(2,3,4-tri-O-benzyl-6-O-
acetoxy-a-D-glucopyranosyl)-L-serine allyl ester (11a)
organic layers were dried (MgSO4), filtered, and concentrated. The
resulting residue was then dissolved in DMF and caesium carbon-
ate (147 mg, 0.45 mmol) and iodomethane (20 mL, 0.30 mmol) were
added. The reaction mixture was stirred at room temperature for
another 2 h, after which TLC analysis showed completion of the
reaction. The solvent was removed under reduced pressure and
EtOAc was added. The organic layer was washed with water and
brine, dried (MgSO4), and concentrated in vacuo. The residue was
purified by silica gel column chromatography to give the esterified
product (117 mg, 0.13 mmol, 88% over two steps). The esterified
product was subjected to a catalytic hydrogenation to give CCL-
34-S1 as a white foam (62 mg, 0.10 mmol, 76%). Rf 0.53 (1 : 9
Compound 11a was obtained in 54% from 11 as white foam. Rf
0.32 (1 : 2 EtOAc–hexane);1H NMR (400 MHz, CDCl3) d 3.47
(1H, dd, J = 9.4, 9.4), 3.53 (1H, dd, J = 3.5, 9.4), 3.82–3.86 (1H,
m), 3.89 (1H, dd, J = 2.9, 10.7 Hz), 3.96 (1H, dd, J = 9.4, 9.4
Hz), 4.09 (1H, dd, J = 3.2, 10.7 Hz), 4.21–4.26 (2H, m), 4.32 (1H,
dd, J = 1.4, 11.8 Hz), 4.39 (2H, d, J = 6.5 Hz), 4.56–4.65 (5H,
m), 4.70 (1H, d, J = 12.2), 4.73 (1H, d, J = 3.5), 4.83 (1H, d, J =
10.8), 4.88 (1H, d, J = 10.8 Hz), 4.99 (1H, d, J = 10.8 Hz), 5.20
(1H, d, J = 10.5 Hz), 5.30 (1H, d, J = 17.2 Hz), 5.83–5.93 (1H,
m), 5.99 (1H, d, J = 8.6 Hz), 7.27–7.39 (19H, m), 7.61 (2H, d, J =
7.4 Hz), 7.75 (1H, d, J = 7.6 Hz);13C (100 MHz, CDCl3) d 20.33,
46.67, 54.22, 62.59, 65.81, 66.78, 69.04, 69.30, 72.46, 74.63, 75.20,
76.82, 79.54, 81.21, 97.75, 118.34, 119.58, 124.73, 126.69, 127.29,
127.38, 127.52, 127.73, 128.02, 128.05, 131.13, 137.43, 137.69,
138.22, 140.85, 143.42, 155.54, 169.20, 170.09; HRMS (ESI) calc.
for C50H52NO11 [M + H]+ 842.3540, found 842.3544.
1
MeOH–CH2Cl2); H (400 MHz, CDCl3/MeOD) d 0.52 (6H, t,
J = 6.8), 0.91–0.93 (32H, m), 1.11–1.17 (2H, m), 1.21–1.27 (2H,
m), 1.88 (2H, t, J = 7.6), 2.82 (2H, t, J = 7.2), 3.35–3.49 (7H,
m), 3.86 (1H, dd, J = 1.6, 3.2), 4.06 (1H, d, J = 1.6), 4.17 (1H,
dd, J = 5.8, 6.6), 4.62 (1H, d, J = 4.0); 13C (100 MHz, CDCl3) d
14.04, 22.62, 25.61, 26.94, 29.31, 29.36, 29.52, 29.62, 31.86, 36.44,
39.73, 51.98, 52.44, 68.17, 68.73, 69.72, 70.48, 70.87, 99.71, 169.51,
173.90; HRMS (ESI) calc. for C33H62N2O9Na [M + Na]+ 653.4353,
found 653.4348.
O-(2,3,4-Tri-O-benzyl-6-hydroxy-a-D-glucopyranosyl)-N-
dodecanoyl-L-serine undecylamide (16)
O-a-D-Glucopyranosyl-N-dodecanoyl-L-serine undecylamide
(CCL-34-S3)
Compound 16 was obtained in 46% from 11a as white foam. Rf
0.32 (1 : 1 EtOAc-hexane); 1H (400 MHz, CDCl3) d 0.83–0.87 (6H,
m), 1.18–1.25 (34H, m), 1.56–1.66 (2H, m), 2.17 (2H, t, J = 7.9 Hz),
2.65–2.73 (1H, m), 3.11–3.18 (1H, m), 3.42 (1H, dd, J = 9.9, 11.1
Hz), 3.55 (1H, dd, J = 9.7, 9.7 Hz), 3.57 (1H, dd, J = 3.6, 9.7 Hz),
3.64–3.70 (2H, m, H-5), 3.77 (1H, d, J = 9.4 Hz), 3.90 (1H, dd, J =
9.7, 9.7 Hz), 3.90 (1H, dd, J = 4.2, 11.1 Hz), 4.33 (1H, ddd, J = 4.2,
6.2, 9.9 Hz), 4.64 (1H, d, J = 11.0 Hz), 4.68 (1H, d, J = 10.8 Hz),
4.79 (1H, d, J = 10.8 Hz), 4.84 (1H, d, J = 11.0 Hz), 4.85 (1H, d, J =
11.0 Hz), 4.90 (1H, d, J = 11.0 Hz), 5.12 (1H, d, J = 3.6 Hz), 6.57
(1H, d, J = 6.2 Hz), 6.94 (1H, t, J = 5.7 Hz), 7.25–7.34 (15H, m); 13C
(100 MHz, CDCl3) d 14.04, 22.60, 25.44, 26.83, 29.26, 29.41, 29.54,
31.82, 36.40, 39.56, 51.14, 61.50, 67.11, 71.58, 74.49, 74.99, 75.57,
77.50, 79.83, 82.07, 97.05, 127.51, 127.65, 127.78, 128.37, 128.39,
128.51, 128.59, 136.98, 137.96, 138.40, 169.21, 173.12; HRMS
(ESI) calc. for C53H81N2O8 [M + H]+ 873.5993, found 873.5991.
Compound CCL-34-S3 was obtained in 83% from 16 as white
foam. Rf 0.23 (1 : 9 MeOH-CH2Cl2); 1H (400 MHz, MeOD) d 0.90
(6H, t, J = 6.7), 1.29 (32H, m), 1.48–1.55 (2H, m), 1.58–1.65 (2H,
m), 2.27 (2H, t, J = 7.5 Hz), 3.14–3.25 (2H, m), 3.29 (1H, dd, J =
9.7 Hz), 3.42 (1H, dd, J = 3.7, 9.7 Hz), 3.53 (1H, ddd, J = 2.2, 5.5,
9.7 Hz), 3.60 (1H, dd, J = 9.7, 9.7 Hz), 3.66 (1H, dd, J = 5.5, 11.8
Hz), 3.74 (1H, dd, J = 6.0, 10.4 Hz), 3.81 (1H, dd, J = 2.2, 11.8 Hz),
3.84 (1H, dd, J = 6.0, 10.4 Hz), 4.53 (1H, dd, J = 6.0, 6.0 Hz), 4.83
(1H, d, J = 3.7 Hz); 13C (100 MHz, MeOD) d 14.48, 23.73, 26.82,
28.02, 30.37, 30.48, 30.66, 30.77, 33.07, 36.84, 40.60, 54.64, 62.56,
68.89, 71.54, 73.35, 74.00, 74.97, 100.81, 171.98, 176.22; HRMS
(ESI) calc. for C32H63N2O8[M + H]+ 603.4584, found 603.4563.
Methyl 1-(N-dodecanoyl undecylcarbamoyl-L-serine)-
a-D-glucopyranosiduronate (CCL-34-S4)
General procedure for the catalytic hydrogenation
Compound CCL-34-S4 was obtained in 67% from 16 as white
1
To a solution of the target compound (0.1 mmol) in a co-solvent
system of CH2Cl2 and methanol (1 : 1, 5 mL), Pd/C (20 mg)
was added. The reaction was shaken under a high pressure of
hydrogen (50 kg cm-2) for 6 h. The reaction mixture was filtered
over a short pad of Celite, and the solid residue was washed
with CH2Cl2/MeOH (1 : 1). The filtrate was concentrated under
reduced pressure, and the resulting residue was purified by silica
gel chromatography to give the product.
foam. Rf 0.37 (1 : 9 MeOH–CH2Cl2); H (400 MHz, CDCl3) d
0.67–0.70 (6H, m), 1.07–1.11 (34H, m), 1.28–1.35 (2H, m), 1.39–
1.46 (2H, m), 2.06 (2H, t, J = 8.0 Hz), 2.95–3.07 (2H, m), 3.35 (1H,
dd, J = 3.6, 9.2 Hz), 3.39 (1H, dd, J = 9.2, 9.6 Hz), 3.46 (1H, dd,
J = 9.2, 9.2 Hz), 3.56–3.58 (2H, m), 3.61 (3H, s), 3.89 (1H, d, J =
9.6 Hz), 4.33 (1H, dd, J = 5.2, 6.4 Hz), 4.74 (1H, d, J = 3.6 Hz);
13C (100 MHz, CDCl3) d 14.10, 22.68, 25.64, 27.00, 29.36, 29.48,
29.56, 29.66, 31.92, 36.52, 39.87, 52.34, 52.70, 69.06, 71.15, 71.36,
71.77, 73.23, 99.58, 169.51, 170.59, 174.18; HRMS (ESI) calc. for
C33H62N2O9Na [M + Na]+ 653.4353, found 653.4355.
Methyl 1-(N-dodecanoyl undecylcarbamoyl-L-serine)-
a-D-galactopyranosiduronate (CCL-34-S1)
O-(2,3-Di-O-para-methoxybenzyl-4,6-O-benzylidene-a-D-
galactopyranosyl)-N-dodecanoyl-L-serine undecylamide (17)
To a vigorously stirred solution of 15 (130 mg, 0.15 mmol) in
CH2Cl2(2 mL), acetone (1 mL), and H2O (1 mL), TEMPO (12 mg,
0.08 mmol) and BAIB (257 mg, 0.38 mmol) were added. After
being stirred at room temperature for 3 h, the reaction mixture
was quenched by addition of a saturated Na2S2O3 solution. The
mixture was then extracted twice with EtOAc and the combined
Compound 17 was obtained in 53% from 8a as white foam. Rf
1
0.40 (1 : 1 EtOAc–hexane); H (400 MHz, CDCl3) d 0.86 (6H, t,
J = 6.5, 7.1 Hz), 1.15–1.23 (34H, m), 1.56 (2H, br s), 2.15 (2H, t,
J = 7.4 Hz), 2.54–2.61 (1H, m), 3.00–3.06 (1H, m), 3.40 (1H, dd,
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J = 10.7,10.7 Hz), 3.62 (1H, br s), 3.77 (3H, s), 3.79 (3H, s), 3.85
(1H, dd, J = 3.1, 10.0 Hz), 3.88 (1H, dd, J = 3.9, 10.7 Hz), 3.96
(1H, dd, J = 1.4, 12.4 Hz), 4.09 (1H, dd, J = 3.5, 10.0 Hz), 4.15
(1H, d, J = 3.1 Hz), 4.21 (1H, dd, J = 1.0, 12.4 Hz), 4.30–4.35
(1H, m), 4.61 (1H, d, J = 10.2 Hz), 4.64 (1H, d, J = 11.8 Hz), 4.67
(1H, d, J = 11.8 Hz), 4.80 (1H, d, J = 10.2 Hz), 5.24 (1H, d, J =
3.5 Hz), 5.44 (1H, s), 6.48 (1H, d, J = 6.2 Hz), 6.82 (2H, d, J =
8.6 Hz), 6.86 (1H, d, J = 8.6 Hz), 7.07 (1H, t, J = 4.8 Hz), 7.23
(2H, d, J = 8.6 Hz), 7.30 (2H, d, J = 8.6 Hz), 7.32–7.39 (5H, m),
7.50 (1H, d, J = 1.4 Hz), 7.52 (1H, d, J = 2.1 Hz);dC (100 MHz,
CDCl3) 14.06, 22.64, 25.52, 26.85, 29.25, 29.32, 29.44, 29.58, 29.62,
31.87, 36.49, 39.42, 51.01, 55.21, 63.08, 67.55, 69.30, 71.29, 74.11,
74.69, 75.23, 76.45, 98.60, 101.12, 113.77, 113.88, 126.33, 128.12,
128.91, 129.05, 129.62, 130.29, 130.47, 137.82, 159.29, 159.60,
169.31, 172.77; HRMS (ESI) calc. for C55H82N2O10Na [M + Na]+
953.5867, found 953.5864.
was purified by silica gel column chromatography to give the azide
(612 mg, 0.70 mmol, 71%). The azide (523 mg, 0.60 mmol) was
catalytically hydrogenated to give 19 as a white foam (435 mg,
1
0.52 mmol, 86%). Rf 0.12 (1 : 9 MeOH–CH2Cl2); H (400 MHz,
CDCl3/MeOD) d 0.77 (3H, t, J = 7.0 Hz), 0.78 (3H, t, J = 7.1
Hz), 1.13–1.24 (34H, m), 1.47–1.54 (2H, m), 2.11 (2H, t, J = 8.0
H), 2.72–2.79 (1H, m), 2.91 (1H, dd, J = 3.8, 13.3 Hz), 2.94–3.01
(1H, m), 3.04 (1H, dd, J = 6.8, 13.3 Hz), 3.49 (1H, dd, J = 9.1, 10.6
Hz), 3.64 (1H, dd, J = 3.2, 10.0), 3.69 (1H, dd, J = 5.1, 10.6 Hz),
3.70–3.72 (7H, m), 3.81 (1H, dd, J = 3.7, 10.0 Hz), 3.94 (1H, d, J =
3.2 Hz), 4.36 (1H, dd, J = 5.1, 9.1 Hz), 4.51 (1H, d, J = 10.8 Hz),
4.52 (1H, d, J = 11.2 Hz), 4.56 (1H, d, J = 11.2 Hz), 4.64 (1H, d, J =
10.8 Hz), 4.93 (1H, d, J = 3.7), 6.74 (2H, d, J = 8.6 Hz), 6.79 (2H,
d, J = 8.6 Hz), 7.16 (2H, d, J = 8.6 Hz), 7.21 (2H, d, J = 8.6 Hz);
13C (100 MHz, CDCl3/MeOD) d 13.33, 22.04, 25.07, 26.33, 28.73,
28.92, 29.01, 31.29, 35.55, 38.95, 41.15, 51.48, 54.48, 66.58, 67.67,
68.21, 71.11,73.35, 74.81, 97.24, 113.21, 113.28, 128.78, 129.21,
129.45, 129.63, 158.81, 159.05, 169.44, 173.75; HRMS (ESI) calc.
for C32H64N3O7[M + H]+ 842.5895, found 842.5900.
O-(2,3-Di-O-para-methoxybenzyl-a-D-galactopyranosyl)-N-
dodecanoyl-L-serine undecylamide (18)
Compound 17 (1.27 g, 1.37 mmol) was dissolved in 10 mL of
MeCN/CH2Cl2/MeOH (15 mL/3 mL/5 mL). Vanadyl triflate
(60 mg, 0.14 mol) was added at ambient temperature and the
resulting mixture was stirred for 6 h. After completion of the
reaction as monitored by TLC, the reaction was quenched by
addition of TEA and the crude product was purified by column
chromatography on silica gel to give 18 as a white foam (60 mg,
0.14 mmol). Rf 0.50 (EtOAc 100%); dH (400 MHz, CDCl3) 0.85
(6H, t, J = 6.5, 7.1 Hz), 1.20–1.25 (34H, m), 1.55–1.62 (2H, m),
2.16 (2H, t, J = 7.9 Hz), 2.64–2.70 (1H, m), 3.05–3.11 (1H, m),
3.42 (1H, dd, J = 9.6, 11.4 Hz), 3.71–3.82 (9H, m), 3.86–3.93 (3H,
m), 4.04 (1H, d, J = 2.3), 4.34 (1H, ddd, J = 4.4, 6.5, 11.4 Hz), 4.60
(1H, d, J = 10.6 Hz), 4.61 (1H, d, J = 11.2 Hz), 4.65 (1H, d, J =
11.2 Hz), 4.75 (1H, d, J = 10.6 Hz), 5.11 (1H, d, J = 3.7), 6.59 (1H,
d, J = 6.5), 6.83 (2H, d, J = 8.6 Hz), 6.88 (2H, d, J = 8.6 Hz), 6.99
(1H, t, J = 5.6 Hz), 7.22 (2H, d, J = 8.6 Hz), 7.27 (2H, d, J = 8.6
Hz); dC (100 MHz, CDCl3) 14.03, 22.60, 25.45, 26.80, 29.28, 29.42,
29.56, 31.83, 36.40, 39.46, 51.20, 55.17, 62.69, 67.76, 68.18, 70.00,
71.84, 74.29, 75.34, 77.67, 98.02, 113.88, 128.42, 128.54, 129.24,
129.48, 129.85, 130.05, 131.96, 132.06, 159.41, 159.57, 169.29,
173.12; HRMS (ESI) calc. for C48H79N2O10[M + H]+ 843.5735,
found 843.5741.
O-(6-Amino-6-deoxy-a-D-galactopyranosyl)-N-dodecanoyl-L-
serine undecylamide (CCL-34-S6)
To a solution of 19 (150 mg, 0.18 mmol) in CH2Cl2 (5 mL),
trifluoroacetic acid (1 mL) was added. The reaction mixture was
stirred at room temperature for 15 min, concentrated, and purified
by column chromatography on silica gel to give CCL-34-S6 as
a white foam (85 mg, 0.14 mmol, 80%). Rf 0.50 (1 : 4 MeOH–
1
CH2Cl2); H (400 MHz, CDCl3/MeOD) d 0.90 (6H, t, J = 6.7
Hz), 1.25–1.35 (32H, m, CH2), 1.49–1.54 (2H, m), 1.58–1.65 (2H,
m), 2.25–2.29 (2H, m), 3.15 (1H, dd, J = 3.6, 13.2), 3.20 (1H, t,
J = 7.4), 3.26 (1H, dd, J = 5.5, 13.2), 3.69 (1H, dd, J = 6.5, 10.1),
3.73 (1H, dd, J = 3.2, 10.1), 3.81 (1H, dd, J = 3.7, 10.1), 3.87–3.91
(2H, m, H-4), 3.95–3.98 (1H, m), 4.54 (1H, dd, J = 6.5, 6.5), 4.93
(1H, d, J = 3.7); 13C (100 MHz, CDCl3/MeOD) d 14.43, 23.61,
26.75, 27.92, 30.23, 30.30, 30.37, 30.44, 30.59, 30.66, 32.95, 36.74,
40.52, 40.64, 41.86, 54.60, 68.21, 68.65, 69.29, 70.52, 71.55, 100.78,
172.08, 172.17, 176.29; HRMS (ESI) calc. for C32H64N3O7[M +
H]+ 602.4744, found 602.4730.
O-(6-[(8-Phenyloctanoyl)amino]-6-deoxy-a-D-galactopyranosyl)-
N-dodecanoyl-L-serine undecylamide (CCL-34-S7)
Compound 19 (68 mg, 0.08 mmol) was dissolved in dry CH2Cl2 (2
mL) and EDCI (31 mg, 0.16 mmol), HOBt (21 mg, 0.16 mmol),
8-phenyloctanoic acid (35 mg, 0.16 mmol), and DIPEA (22 mL,
0.16 mmol) were added. The reaction mixture was stirred at room
temperature for 3 h. The reaction mixture was diluted with CH2Cl2
and washed with 10% aqueous HCl, saturated aqueous NaHCO3,
and brine. The organic layer was dried over MgSO4, concentrated,
and purified by flash silica gel column chromatography to give the
product of amidation. Trifluoroacetic acid (1 mL) was added to a
solution of this product in CH2Cl2 (5 mL). The reaction mixture
was stirred at room temperature for 15 min, concentrated, and
purified by column chromatography on silica gel to give CCL-34-
S7 as a white foam (47 mg, 0.06 mmol, 73% for two steps). Rf 0.40
(1 : 9 MeOH–CH2Cl2); 1H (400 MHz, CDCl3/MeOD) d 0.54 (6H,
t, J = 6.7, 7.0 Hz), 0.92–1.00 (38H, m), 1.12–1.19 (2H, m), 1.22–
1.30 (6H, m), 1.85–1.93 (4H, m), 2.25 (2H, t, J = 7.7), 2.78–2.95
O-(6-Amino-6-deoxy-2,3-di-O-para-methoxybenzyl-a-D-
galactopyranosyl)-N-dodecanoyl-L-serine undecylamide (19)
To a solution of 18 (0.84 g, 0.99 mmol) in pyridine (10 mL), TsCl
(570 mg, 2.99 mmol) was added at 0 ◦C. After being stirred at the
same temperature for 10 min, the reaction was allowed to warm to
room temperature and was stirred for another 12 h. The reaction
was quenched with methanol and concentrated in vacuo. The
reaction mixture was dissolved in EtOAc and washed with water
and brine. The organic layer was dried (MgSO4) and concentrated
under reduced pressure to give the tosylated crude product. To a
solution of the tosylated product in DMF (10 mL), sodium azide
(329 mg, 5.01 mmol) was added and the resulting mixture was
stirred at 80 ◦C for 12 h. The reaction mixture was concentrated,
dissolved in EtOAc, and washed with water and brine. The organic
layer was dried (MgSO4) and concentrated in vacuo. The residue
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(3H, m), 3.16–3.23 (1H, m), 3.32–3.46 (6H, m), 4.16–4.20 (1H,
m), 4.51 (1H, d, J = 3.8), 6.79–6.82 (3H, m), 6.89–6.93 (2H, m),
7.38 (1H, d, J = 7.8), 7.46 (1H, t, J = 5.6), 7.55 (1H, t, J = 5.4);
13C (100 MHz, CDCl3/MeOD) d 13.17, 22.02, 25.09, 25.24, 26.33,
28.51, 28.63, 28.72, 28.91, 29.00, 30.89, 31.30, 35.27, 35.47, 35.57,
39.02, 39.10, 52.31, 66.92, 68.11, 68.59, 68.71, 69.24, 98.93, 124.93,
127.56, 127.70, 142.10, 169.97, 174.23, 175.18; HRMS (ESI) calc.
for C46H81N3O8Na [M + Na]+ 826.5921, found 826.5903.
S-p-Tolyl 4-O-allyl-2,3,6-tri-O-benzyl-b-D-thiogalactopyranoside
(12)
To a solution of S-p-tolyl 4,6-O-benzylidene-2,3-di-O-benzyl-b-
D-thiogalactopyranoside (0.59 g, 1.07 mmol) in THF (22 mL),
Me3NBH3 (0.32 g, 4.39 mmol), AlCl3 (0.86 g, 6.45 mmol), and
H2O (0.04 mL, 2.14 mmol) were added at room temperature. After
being stirred for 4 h, the reaction mixture was quenched by adding
saturated aqueous HCl and extracted with EtOAc. The combined
organic layers were dried over MgSO4, filtered, concentrated, and
purified by column chromatography to give the selectively reduced
product (corresponding to the 6-O-benzyl derivative) that was then
dissolved in DMF (2.7 mL). NaH (23.4 mg, 1.56 mmol) and allyl
O-(6-[(2-Hydroxydodecanoyl)amino]-6-deoxy-a-D-galactopyra-
nosyl)-N-dodecanoyl-L-serine undecylamide (CCL-34-S8)
Compound CCL-34-S8 was obtained from 19 (68 mg, 0.08 mmol)
as a white foam (44 mg, 0.05 mmol, 69%). Rf 0.33 (1 : 9 MeOH–
CH2Cl2); 1H (400 MHz, CDCl3/MeOD) d 0.52 (9H, t, J = 6.7, 7.0
Hz), 0.82–1.51 (50H, m), 1.81–1.86 (2H, m), 2.69–2.82 (2H, m),
2.92–2.96 (1H, m), 3.09–3.16 (1H, m), 3.23–3.38 (6H, m), 3.60–
3.66 (1H, m, H-5¢), 4.08–4.11 (1H, m), 4.42–4.44 (1H, m); 13C
(100 MHz, CDCl3/MeOD) d 13.22, 22.05, 24.67, 24.73, 25.12,
26.36, 28.63, 28.76, 28.88, 28.95, 29.04, 31.33, 33.93, 34.08, 35.51,
38.80, 39.14, 52.32, 52.40, 66.70, 66.75, 68.14, 68.60, 68.85, 69.05,
69.34, 69.40, 71.27, 77.19, 98.80, 98.87, 169.92, 169.98, 174.26,
174.30, 176.07, 176.31; HRMS (ESI) calc. for C44H85N3O9Na [M +
Na]+ 822.6184, found 822.6169.
◦
bromide (0.076 ml, 1.25 mmol) were added at 0 C. After being
stirred for 3 h, the reaction mixture was quenched by the addition
of MeOH, extracted with ethyl acetate, and washed with brine.
The combined organic layers were dried over MgSO4, filtered, and
purified by column chromatography to give compound12 (0.44 g,
91%). Rf 0.4 (1 : 4 EtOAc–Hexane); 1H (400 MHz, CDCl3) d 2.29
(3H, s), 3.52–3.57 (2H, m), 3.66 (1H, dd, J = 5.5, 9.2 Hz), 3.72
(1H, dd, J = 9.2, 9.6 Hz), 3.83 (1H, t, J = 9.7 Hz), 3.87 (1H, d, J =
2.6 Hz), 4.10 (1H, dd, J = 5.9, 12.8 Hz), 4.38 (1H, dd, J = 5.3, 12.8
Hz), 4.49 (1H, d, J = 11.6 Hz) 4.52 (1H, d, J = 11.6 H), 4.57 (1H, d,
J = 9.7), 4.67 (1H, d, J = 11.6 Hz), 4.72 (1H, d, J = 11.6 Hz), 4.73
(1H, d, J = 10.2 Hz), 4.80 (1H, d, J = 10.2 Hz), 5.13 (1H, dd, J =
1.6, 10.3 Hz), 5.23 (1H, dd, J = 1.6, 17.2 Hz), 5.89 (1H, m), 7.04
(2H, d, J = 7.9 Hz), 7.27–7.40 (15H, m), 7.46 (2H, d, J = 6.4 Hz);
13C (100 MHz, CDCl3) d 21.01, 68.64, 72.57, 73.20, 73.57, 75.57,
77.32, 77.43, 83.91, 88.22, 116.43, 127.54, 127.61, 127.72, 127.85,
128.23, 128.34, 129.48, 130.36, 132.11, 135.35, 137.16, 137.85,
138.15, 138.35, 138.35. HRMS (ESI) calc. for C37H40O5SNa [M +
Na+]: 619.2494, found 619.2498.
O-(6-Butyrylamino-6-deoxy-a-D-galactopyranosyl)-N-
dodecanoyl-L-serine undecylamide (CCL-34-S9)
Compound CCL-34-S9 was obtained from 19 (35 mg, 0.04 mmol)
as a white foam (22 mg, 0.03 mmol, 78%). Rf 0.37 (1 : 9 MeOH–
1
CH2Cl2); H (400 MHz, CDCl3/MeOD) d 0.52 (6H, t, J = 6.7,
7.0 Hz), 0.58 (3H, t, J = 7.4), 0.88–0.97 (32H, m), 1.11–1.17 (2H,
m), 1.22–1.31 (4H, m), 1.83 (2H, t, J = 7.4), 1.89 (2H, t, J = 7.6),
2.77–2.92 (3H, m), 3.15–3.20 (1H, m), 3.30–3.44 (6H, m), 4.15
(1H, dd, J = 5.9, 5.9), 4.49 (1H, d, J = 3.7), 7.39 (1H, d, J =
7.4), 7.47 (1H, t, J = 5.6), 7.53 (1H, t, J = 5.4); 13C (100 MHz,
CDCl3/MeOD) d 12.46, 12.80, 18.42, 21.80, 24.90, 26.11,28.46,
28.51, 28.70, 28.79, 31.10, 35.16, 37.17, 38.82, 38.95, 39.05, 39.17,
52.26, 66.71, 67.96, 68.64, 68.83, 69.12, 98.88, 169.87, 169.96,
174.13, 174.75; HRMS (ESI) calc. for C36H69N3O8Na [M + Na]+
694.4982, found 694.4989.
N-(9-Fluorenylmethoxycarbonyl)-O-(4-O-allyl-2,3,6-tri-O-
benzyl-a-D-galactopyranosyl)-L-serine allyl ester (12a)
Compound 12a was obtained from 12 in 66% as white foam. Rf
0.38 (1 : 2 EtOAc–hexane); 1H (400 MHz, CDCl3) d 3.58 (2H, d,
J = 6.5 Hz), 3.76–3.82 (3H, m), 3.91–3.95 (2H, m), 4.02 (1H, dd,
J = 6.2, 12.4 Hz), 4.13 (1H, t, J = 7.2 Hz), 4.18 (1H, dd, J = 3.6,
11.2 Hz), 4.27–4.35 (3H, m), 4.42 (1H, d, J = 12.0 Hz), 4.49–4.58
(5H, m), 4.65–4.74 (4H, m), 5.06–5.26 (4H, m), 5.79–5.85 (2H,
m), 6.21 (1H, d, J = 8.6 Hz), 7.20–7.34 (19H, m), 7.53 (2H, d, J =
7.4 Hz), 7.70 (2H, d, J = 7.6 Hz); 13C (100 MHz, CDCl3) d 47.05,
54.71, 66.05, 67.04, 68.72, 69.86, 70.31, 72.86, 73.29, 73.40, 73.93,
74.68, 76.42, 78.36, 99.57, 116.93, 118.56, 119.86, 125.11, 126.99,
127.39, 127.44, 127.58, 127.70, 128.-8, 128.28, 131.54, 135.21,
137.86, 138.50, 138.57, 141.20, 143.79, 143.84, 156.05, 169.71;
HRMS (ESI) calc. for C51H53NO10Na [M + Na]+ 862.3567, found
862.3566.
O-(6-[5-(2-Oxoperhydrothieno[3,4-d]imidazol-4-yl)pentanoyl-
amino]-6-deoxy-a-D-galactopyranosyl)-N-dodecanoyl-L-serine
undecylamide (CCL-34-S10)
Compound CCL-34-S10 was obtained from 19 (68 mg, 0.11 mmol)
as a white foam (65 mg, 0.08 mmol, 71%). Rf 0.13 (1 : 9 MeOH–
1
CH2Cl2); H (400 MHz, CDCl3/MeOD) d 0.86 (6H, t, J = 6.7,
7.0 Hz), 1.22–1.28 (32H, m), 1.41–1.47 (4H, m), 1.56–1.69 (6H,
m), 2.22–2.27 (4H, m), 2.71 (1H, d, J = 12.8), 2.91 (1H, dd,
J = 5.0, 12.8), 3.11–3.20 (3H, m), 3.26–3.30 (1H, m), 3.48 (1H,
dd, J = 6.6, 13.7), 3.65–3.79 (6H, m), 4.31 (1H, dd, J = 4.5,
7.8), 4.48–4.53 (2H, m), 4.84 (1H, d, J = 3.8); 13C (100 MHz,
CDCl3/MeOD) d 13.04, 21.95, 24.87, 25.06, 26.28, 27.54, 27.82,
28.54, 28.66, 28.73, 28.87, 28.94, 31.24, 34.91, 35.36, 38.99, 39.05,
39.49, 52.49, 55.07, 59.73, 61.50, 66.75, 68.05, 68.49, 68.82, 69.17,
77.20, 98.86, 114.62, 117.52, 170.08, 174.38, 174.84; HRMS (ESI)
calc. for C42H77N5O9SNa [M + Na]+ 850.5340, found 850.5341.
O-(4-O-Propyl)-a-D-galactopyranosyl)-N-dodecanoyl-L-serine
undecylamide (CCL-34-S11)
Compound CCL-34-S11 was obtained from 12a in 43% for five
steps as a white foam. Rf 0.15 (4 : 1 EtOAc–CH2Cl2); 1H (400 MHz,
CDCl3) d 0.87–0.92 (9H, m), 1.16–1.33 (32H, m), 1.46 (2H, m),
1.56–1.60 (4H, m), 2.20 (2H, t, J = 7.5 Hz), 2.51 (1H, d, J = 6.2 Hz),
2.84 (1H, bs), 3.04–3.11 (2H, m), 3.38 (1H, bs), 3.48–3.54 (1H, m),
3.63–3.88 (9H, m), 4.61 (1H, dd, J = 6.8, 12.1 Hz), 5.03 (1H, d,
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J 3.4 Hz), 6.84 (1H, d, J = 7.5 Hz), 7.19 (1H, bs); 13C (100 MHz,
CDCl3) d 10.43, 14.05, 22.63, 23.35, 25.62, 26.99, 29.32, 29.61,
31.87, 36.42, 39.85, 52.31, 62.45, 68.53, 69.85, 71.17, 71.99, 75.62,
77.32, 99.48, 169.98, 174.08; HRMS (ESI) calc. for C35H68N2O8Na
[M + Na]+ 667.4873, found 667.4868.
d, J = 7.4 Hz), 7.74 (2H, d, J = 7.7 Hz); 13C (100 MHz, CDCl3) d
46.95, 54.65, 55.08, 63.28, 66.10, 67.05, 69.15, 70.32, 71.10, 73.15,
74.43, 74.85, 75.12, 100.20, 100.82, 113.66, 116.76, 118.59, 119.88,
124.90, 124.98, 126.13, 126.95, 126.98, 127.60, 127.96, 128.74,
129.40, 130.47, 131.39, 135.05, 137.65, 141.14, 143.62, 155.96,
159.15, 169.80; HRMS (ESI) calc. for C45H47NO11Na [M + Na]+
800.3047, found 800.3046.
S-p-Tolyl 3-O-allyl-2-O-para-methoxybenzyl-
4,6-O-benzylidene-b-D-thiogalactopyranoside (13)
O-(3-O-Allyl-a-D-galactopyranosyl)-N-dodecanoyl-L-serine
A
suspension
of
S-p-tolyl
4,6-O-benzylidene-b-D-
undecylamide (CCL-34-S12)
thiogalactopyranoside (1 g, 2.68 mmol) and dibutyltin oxide
(1 g, 4.00 mmol) in toluene (20 mL) was heated to reflux in a
Dean–Stark apparatus for 4 h, after which most of the solvent
was distilled off. The reaction mixture was cooled to room
temperature, and the residual solvent was evaporated under
reduced pressure. CsF (820 mg, 5.36 mmol), allyl bromide
(0.46 ml, 5.36 mmol), tetrabutylammonium iodide (2 g, 5.36
mmol), and DMF (13 mL) were then added. The reaction mixture
was heated to reflux overnight. It was then diluted with ethyl
acetate, washed (sat. aq. NaHCO3, brine), dried (MgSO4), and
concentrated. Purification by column chromatography gave the
allylated product (981 mg, 2.37 mmol, 88%) as a white foam.
To a solution of the allylated product and TBAI in DMF, NaH
(189 mg, 4.72 mmol) was added at 0 ◦C. The resulting mixture was
stirred for 10 min in an ice–water bath and then p-methoxybenzyl
chloride (0.5 mL, 3.54 mmol) was added. The reaction mixture
was warmed to room temperature and stirred for another 4 h until
TLC analysis showed completion of the reaction. The mixture was
then quenched with methanol, concentrated in vacuo, dissolved
in EtOAc, and washed with water and brine. The organic layer
was dried (MgSO4), filtered, and concentrated. The resulting
residue was purified by silica gel column chromatography to give
Compound CCL-34-S12 was obtained from 13a in 34% for five
steps as white foam. Rf 0.3 (1 : 20 MeOH–CH2Cl2); 1H (400 MHz,
CDCl3) d 0.85 (7H, m), 1.00 (1H, t, J = 7.2 Hz), 1.10–1.37 (37H,
m), 1.37–1.70 (5H, m), 2.19 (3H, dd, J = 8.0, 7.2 Hz), 3.09–3.37
(4H, m), 3.52 (1H, d, J = 9.6 Hz), 3.62 (1H, dd, J = 10.4, 9.2 Hz),
3.72–3.95 (5H, m), 3.96–4.26 (5H, m), 4.56–4.59 (1H, m), 5.11 (1H,
d, J = 3.7 Hz), 5.18 (1H, dd, J = 10.4, 1.7 Hz), 5.27 (1H, dd, J =
17.6, 1.7 Hz), 5.91 (1H, dddd, J = 5.2, 5.2, 5.6, 5.6 Hz), 6.90 (1H),
7.55 (1H); 13C (100 MHz, CDCl3) d 14.10, 22.67, 24.22, 25.60,
26.98, 29.35, 29.49, 29.52, 29.62, 31.90, 36.54, 39.83, 51.73, 62.94,
68.00, 68.27, 70.39, 70.86, 99.38, 117.62, 134.55, 169.64, 173.81;
HRMS (ESI) calc. for C35H66N2O8Na [M + Na]+ 665.4717, found
665.4720.
N-(9-Fluorenylmethoxycarbonyl)-O-(2,3,4-tri-O-benzyl-a-L-
rhamnopyranosyl)-L-serine allyl ester (14a)
Compound 14a was obtained from 14 in 56% yield for 2 steps as
white foam. Rf 0.31 (1 : 2 EtOAc–hexane); 1H (400 MHz, CDCl3)
d 1.07 (3H, d, J = 6.5), 3.56–3.59 (2H, m), 3.74 (1H, dd, J = 6.4,
12.9), 3.83 (1H, dd, J = 2.6, 10.1), 4.01 (1H, dd, J = 3.6, 10.1), 4.20–
4.23 (2H, m), 4.35 (1H, dd, J = 7.2, 10.5), 4.47 (1H, dd, J = 6.9,
10.5), 4.53–4.58 (1H, m), 4.61–4.64 (4H, m), 4.67 (1H, d, J = 3.6),
4.72 (1H, d, J = 11.8), 4.79 (1H, d, J = 12.1), 4.83 (1H, d, J = 11.8),
4.96 (1H, d, J = 11.6), 5.18 (1H, dd, J = 1.2, 10.3), 5.29 (1H, dd, J =
1.2, 17.2), 5.86 (1H, ddd, J = 9.0, 10.3, 17.2), 6.18 (1H, d, J = 9.0),
1
13 (853 mg, 2.05 mmol, 87%). Rf 0.25 (1 : 3 EtOAc–hexane); H
(400 MHz, CDCl3) d 2.30 (3H, s), 3.42 (1H, s), 3.52 (1H, dd, J =
9.2, 3.6 Hz), 3.78 (1H, dd, J = 9.6, 9.2 Hz), 3.79 (3H, s), 4.00
(1H, dd, J = 12.4, 1.6 Hz), 4.17–4.22 (3H, m), 4.37 (1H, dd, J =
12.4, 1.6 Hz), 4.55 (1H, d, J = 9.6 Hz), 4.58 (1H, d, J = 10.6 Hz),
4.61 (1H, d, J = 10.6 Hz), 5.17 (1H, dddd, J = 10.5, 1.7, 1.4, 1.4
Hz), 5.29 (1H, dddd, J = 17.2, 1.7, 1.7, 1.7 Hz), 5.52 (1H, s),
5.87–5.97 (1H, m), 6.85–6.89 (2H, m), 7.00 (2H, d, J = 8.0 Hz),
7.35–7.37 (5H, m), 7.47–7.49 (2H, m), d 7.60 (2H, d, J = 8.4 Hz);
13C (100 MHz, CDCl3) d 20.88, 54.98, 69.12, 69.44, 70.81, 73.55,
74.71, 74.91, 76.68, 77.00, 77.32, 80.94, 86.33, 100.91, 113.47,
117.03, 126.42, 127.80, 128.21, 128.67, 128.71, 129.38, 129.53,
130.57, 132.94, 134.64, 137.23, 137.76; HRMS (ESI) calc. for
C31H34O6SNa [M + Na]+ 557.1974, found 557.1981.
7.19–7.40 (19H, m), 7.59 (2H, t, J = 7.0), 7.72 (2H, d, J = 7.6); 13
C
(100 MHz, CDCl3) d 16.30, 46.85, 54.28, 65.63, 66.57, 66.76, 68.80,
72.75, 73.04, 74.55, 76.01, 78.63, 98.82, 118.16, 119.67, 124.74,
124.81, 126.77, 127.25, 127.33, 127.39, 127.58, 127.72, 127.91,
128.05, 128.09, 131.36, 138.22, 138.45, 140.95, 143.44, 143.66,
156.01, 169.59; HRMS (ESI) calc. for C48H49NO9Na [M + Na]+
806.3305, found 806.3300.
O-a-L-Rhamnopyranosyl-N-dodecanoyl-L-serine undecylamide
(CCL-34-S13)
Compound CCL-34-S13 was obtained from 14a in 57% yield for 5
steps as white foam. Rf 0.38 (1 : 9 MeOH–CH2Cl2); 1H (400 MHz,
MeOD) d 0.90 (6H, t, J = 6.7), 1.19 (3H, d, J = 6.5), 1.28–1.36
(32H, m), 1.45–1.52 (2H, m), 1.61–1.67 (2H, m), 2.31 (2H, t, J =
7.5), 3.16–3.21 (2H, m), 3.48 (1H, dd, J = 3.3, 9.7), 3.62 (1H, d,
J = 3.2), 3.68 (1H, dd, J = 3.2, 10.0), 3.77 (1H, dd, J = 3.7, 10.0),
3.80–3.85 (1H, m), 4.11 (1H, dd, J = 3.1, 9.7), 4.51–4.55 (1H, m),
4.74 (1H, d, J = 3.7), 7.88 (1H, t, J = 5.5), 8.46 (1H, d, J = 8.0);
13C (100 MHz, CDCl3/MeOD) d 13.07, 15.12, 21.85, 24.93, 26.18,
28.56, 28.60, 28.68, 28.85, 31.13, 35.15, 38.83, 38.96, 52.31, 65.71,
67.10, 67.96, 69.70, 71.39, 98.15, 169.91, 173.91; HRMS (ESI)
calc. for C32H62N2O7Na [M + Na]+ 609.4455, found 609.4462.
N-(9-Fluorenylmethoxycarbonyl)-O-(2-O-para-methoxybenzyl-3-
O-allyl-4,6-O-benzylidene-a-D-galactopyranosyl)-L-serine allyl
ester (13a)
Compound 13a was obtained from 13 in 57% as white foam. Rf
0.13 (1 : 3 EtOAc–hexane); 1H (400 MHz, CDCl3) d 3.68 (1H, s),
3.75 (3H, s), 3.78–3.86 (2H, m), 3.97 (1H, dd, J = 14.3, 14.3 Hz),
3.99 (1H, d, J = 7.4 Hz), 4.14–4.28 (6H, m), 4.30–4.45 (2H, m),
4.52 (1H, d, J = 11.6 Hz), 4.61 (2H, m), 4.72 (1H, d, J = 11.6 Hz),
4.79 (1H, d, J = 3.0 Hz), 5.15–5.23 (2H, m), 5.27–5.34 (2H, m),
5.51 (1H, s), 5.81–6.00 (2H, m), 6.16 (1H, NH), 6.84 (2H, d, J =
8.0 Hz), 7.24–7.39 (10H, m), 7.50 (2H, d, J = 6.8 Hz), 7.58 (2H,
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˚
O-b-D-Galactopyranosyl-N-dodecanoyl-L-serine undecylamide
(b-CCL-34)
and activated 4 A MS (2.18 g) in CH2Cl2 (10 mL). After being
stirred for 30 min at -40 ◦C, the reaction mixture was warmed to
room temperature and stirred for another 12 h. It was then filtered
over a pad of Celite and the filtrate was concentrated. The obtained
residue was purified by column chromatography to give the pure
product (1.3 g, 86%). Rf 0.25 (1 : 3 EtOAc–hexane); 1H (400 MHz,
CDCl3/MeOD) d 1.98 (3H, s), 2.00 (3H, s), 2.04 (3H, s), 3.60 (1H,
dd, J = 8.7, 10.2 Hz), 3.73 (1H, m), 3.91 (1H, dd, J = 3.6, 10.3 Hz),
4.09 (1H, dd, J = 2.0, 12.4 Hz), 4.18–4.33 (4H, m), 4.44 (1H, dd,
J = 5.9, 9.3 Hz), 4.49 (1H, t, J = 3.6 Hz), 4.53 (1H, d, J = 12.1 Hz),
4.58–4.71 (3H, m), 4.75 (1H, d, J = 12.1 Hz), 4.98 (1H, t, J = 10.2
Hz), 5.21 (1H, dd, J = 1.1, 10.5 Hz), 5.23 (1H, t, J = 10.2 Hz), 5.32
(1H, dd, J = 1.1, 17.2 Hz), 5.86 (1H, ddd, J = 8.0, 10.5, 17.2 Hz).
7.29–7.33 (2H, m), 7.38 (2H, t, J = 7.4 Hz), 7.63 (2H, t, J = 7.0 Hz),
7.75 (2H, d, J = 7.5 Hz); 13C (100 MHz, CDCl3) d 13.76, 20.16,
20.24, 20.59, 29.21, 46.62, 53.75, 55.38, 60.03, 61.67, 65.79, 66.96,
68.35, 68.49, 71.31, 71.66, 73.89, 95.19, 100.06, 118.13, 119.66,
124.74, 124.90, 126.77, 127.41, 131.13, 140.83, 143.19, 143.49,
154.10, 155.89, 169.03, 169.10, 170.11, 170.25; HRMS (ESI) calc.
for C36H39N2O14NaCl3 [M + Na]+ 851.1365, found 851.1374.
Compound b-CCL-34 was obtained from 5 in 43% yield for 5
steps as white foam. Rf 0.25 (1 : 9 MeOH–CH2Cl2); 1H (400 MHz,
CDCl3/MeOD) d 0.62 (3H, t, J = 6.6 Hz), 0.62 (3H, t, J = 7.0 Hz),
0.95–1.09 (32H, br s), 1.21–1.29 (2H, m), 1.32–1.41 (2H, m), 2.00
(2H, t, J = 7.6 Hz), 2.93 (2H, dd, J = 7.3, 13.4 Hz), 3.21–3.32 (3H,
m), 3.44 (1H, dd, J = 5.2, 10.3 Hz), 3.47 (1H, dd, J = 4.7, 13.2 Hz),
3.54–3.63 (2H, m), 3.84 (1H, dd, J = 5.2, 10.3 Hz), 3.97 (1H, d, J =
7.3 Hz), 4.34 (1H, t, J = 5.2 Hz); 13C (100 MHz, CDCl3/MeOD) d
13.29, 22.08, 25.12, 26.35, 28.61, 28.77, 28.84, 28.97, 29.06, 31.35,
35.56, 39.15, 52.68, 60.97, 68.47, 69.00, 70.65, 72.89, 74.80, 103.40,
169.91, 174.29; HRMS (ESI) calc. for C32H62N2O8Na [M + Na]+
625.4404, found 625.4401.
O-a-D-Glucopyranosyl-N-dodecanoyl-L-serine undecylamide
(CCL-34-S14)
Compound CCL-34-S14 was obtained from 20 in 55% yield for 5
steps as white foam. Rf 0.23 (1 : 9 MeOH–CH2Cl2); 1H (400 MHz,
MeOD) d 0.90 (6H, t, J = 6.7, 7.0), 1.29 (32H, br s), 1.48–1.55 (2H,
m), 1.58–1.65 (2H, m), 2.27 (2H, t, J = 7.5 Hz), 3.14–3.25 (2H,
m), 3.29 (1H, dd, J = 9.7 Hz), 3.42 (1H, dd, J = 3.7, 9.7 Hz), 3.53
(1H, ddd, J = 2.2, 5.5, 9.7 Hz), 3.60 (1H, dd, J = 9.7, 9.7 Hz), 3.66
(1H, dd, J = 5.5, 11.8 Hz), 3.74 (1H, dd, J = 6.0, 10.4 Hz), 3.81
(1H, dd, J = 2.2, 11.8 Hz), 3.84 (1H, dd, J = 6.0, 10.4 Hz), 4.53
(1H, dd, J = 6.0, 6.0 Hz), 4.83 (1H, d, J = 3.7 Hz); 13C (100 MHz,
MeOD) d 14.48, 23.73, 26.82, 28.02, 30.37, 30.48, 30.66, 30.77,
33.07, 36.84, 40.60, 54.64, 62.56, 68.89, 71.54, 73.35, 74.00, 74.97,
100.81, 171.98, 176.22; HRMS (ESI) calc. for C32H63N2O8[M +
H]+ 603.4584, found 603.4563.
N-(2,2,2-Trichloroethyloxycarbonyl-2-amino-2-deoxy)-O-(3,4,6-
tri-O-acetoxy-b-D-glucopyranosyl)-N-dodecanoyl-L-serine
undecylamide (25)
Rf 0.28 (1 : 3 EtOAc–CH2Cl2); 1H (400 MHz, CDCl3/MeOD,
mixture of two conformation isomers) d 0.85 (6H, t, J = 6.7,
7.0 Hz), 1.18–1.35 (32H, m), 1.46–1.50 (2H, m), 1.56–1.60 (2H,
m), 1.97 and 1.98 (3H, s), 2.00 and 2.01 (3H, s), 2.06 and 2.07
(3H, s), 2.22 (2H, t, J = 7.4 Hz), 3.07–3.25 (2H, m), 3.62 and
3.66 (1H, dd, J = 8.4, 10.5 and 8.4, 10.5 Hz), 3.70–3.82 (2H, m),
3.85 and 3.89 (1H, dd, J = 5.5, 10.1 and 6.0, 10.6 Hz), 4.10 and
4.13 (1H, dd, J = 2.2, 14.2 and 2.0, 14.2 Hz), 4.24–4.34 (1H, m),
4.45 and 4.53 (1H, t, J = 5.5 and 6.0 Hz), 4.54 (1H, d, J = 8.4
Hz), 4.57 and 4.58 (1H, d, J = 12.12 and 12.12 Hz), 4.82 and 4.84
(1H, d, J = 12.12 and 12.04 Hz), 4.98 and 4.99 (1H, t, J = 9.8
and 9.8 Hz), 5.18 and 5.22 (1H, dd, J = 9.8, 10.5 and 9.8, 10.5
Hz); 13C (100 MHz, CDCl3/MeOD) d 14.50, 20.92, 21.03, 23.31,
26.32, 27.59, 29.85, 30.01, 30.06, 30.20, 30.29, 32.59, 36.76, 53.32,
53.51, 56.55, 62.73, 69.51, 69.73, 70.16, 72.40, 72.54, 73.10, 73.23,
75.06, 96.35, 96.44, 102.03, 155.93, 156.31, 170.65, 170.73, 171.36,
171.88, 175.38; HRMS (ESI) calc. for C41H70N3O12NaCl3 [M +
Na]+ 924.3923, found 924.3916.
O-a-D-Mannopyranosyl-N-dodecanoyl-L-serine undecylamide
(CCL-34-S15)
Compound CCL-34-S15 was obtained from 22 in 51% yield
1
for four steps as white solid. Rf 0.25 (1 : 9 MeOH–CH2Cl2); H
(400 MHz, CDCl3/MeOD) d 0.61 (6H, t, J = 6.7, 7.0), 0.96–
1.15 (32H, m, CH2), 1.19–1.26 (2H, m), 1.31–1.37 (2H, m), 1.98
(2H, t, J = 7.6), 2.85–2.98 (2H, m), 3.21–3.25 (1H, m), 3.37–3.58
(7H, m), 4.31 (1H, dd, J = 5.7, 5.7), 4.51 (1H, d, J = 1.1); 13C
(100 MHz, CDCl3/MeOD) d 13.34, 22.11,25.20, 26.41, 28.81,
28.87, 29.01, 29.09, 31.38, 35.59, 39.15, 52.48, 61.05, 66.63, 67.04,
70.03, 70.78, 72.61, 100.16, 169.72, 174.24; HRMS (ESI) calc. for
C32H62N2O8Na [M + Na]+ 625.4404, found 625.4401.
N-(2-Amino-2-deoxy)-O-(3,4,6-tri-O-acetoxy-b-D-
glucopyranosyl)-N-dodecanoyl-L-serine undecylamide (26)
N-(9-Fluorenylmethoxycarbonyl)-N-(2,2,2-
trichloroethyloxycarbonyl-2-amino-2-deoxy)-O-(3,4,6-tri-O-
acetoxy-b-D-glucopyranosyl)-L-serine allyl ester (24)
A solution of 25 (240 mg, 0.27 mmol) and Zn (0.7 g, 10.70 mmol) in
tetrahydrofuran (4.0 mL) was stirred for 4 h at room temperature
and filtered over Celite. Tetrahydrofuran was removed with a
water pump and the resulting residue was purified by column
chromatography to the yield the desired product (186 mg, 95%).
Rf 0.06 (EtOAc 100%); 1H (400 MHz, CDCl3/MeOD, mixture of
two conformation isomer) d 0.89 (6H, t, J = 6.6, 7.0 Hz), 1.21–
1.37 (32H, m), 1.46–1.53 (2H, m), 1.60–1.63 (2H, m), 1.98 and
2.00 (3H, s), 2.05 (3H, s), 2.06 (3H, s), 2.26–2.30 (2H, m), 2.87
(1H, dd, J = 8.3, 9.7 Hz), 3.18–3.21 (2H, m), 3.80–3.84 (1H, m),
3.82 and 3.89 (1H, dd, J = 5.0, 10.3 and 5.4, 10.3 Hz), 4.01 and
4.08 (1H, dd, J = 5.2, 10.3 and 5.0, 10.3 Hz), 4.09–4.13 (1H, m),
Compound 23 (2.9 g, 5.55 mmol) was dissolved in HBr/HOAc
(14.5 ml) at 0 ◦C and the resulting solution was stirred for 3 h in an
ice–water bath. The reaction mixture was quenched by addition
of saturated aqueous NaHCO3 and extracted with EtOAc. The
combined organic layers were dried over MgSO4, filtered, and
concentrated under reduced pressure. The residue was dried under
vacuum for 10 min and then redissolved in CH2Cl2 (5.0 mL). The
obtained solution was slowly added to a cooled solution (-4 ◦C) of
Fmoc-Ser-OAll (1.36 g, 3.70 mmol), AgOTf (1.56 g, 6.08 mmol),
2502 | Org. Biomol. Chem., 2011, 9, 2492–2504
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4.30 and 4.31 (1H, dd, J = 3.9, 8.4 and 4.5, 12.4 Hz), 4.49 and 4.54
(1H, d, J = 8.1 and 8.3 Hz), 4.59 (1H, t, J = 5.2 Hz), 4.93 and 4.95
(1H, t, J = 9.7 and 9.6 Hz), 5.10 and 5.12 (1H, dd, J = 9.7, 11.4
and 9.6, 10.1 Hz); 13C (100 MHz, CDCl3/MeOD) d 14.75, 21.04,
21.06, 23.57, 26.53, 27.71, 30.06, 30.16, 30.24, 30.37, 30.44, 32.74,
36.84, 40.43, 53.00, 54.06, 56.66, 57.94, 62.98, 63.39, 69.86, 70.14,
70.26, 70.68, 71.88, 71.94, 72.75, 72.84, 73.14, 73.83, 74.07, 75.42,
103.68, 104.05, 170.97, 171.05, 171.22, 171.58, 172.04, 172.14,
173.17, 175.76; HRMS (ESI) calc. for C38H69N3O10Na [M + Na]+
750.4881, found 750.4882.
N-(2-Dodecanoyl-2-amino-2-deoxy)-O-b-D-glucopyranosyl-N-
dodecanoyl-L-serine undecylamide (CCL-34-S17)
NEt3 (0.02 mL, 0.16 mmol) was added at room temperature to
a solution of 26 (56 mg, 0.08 mmol), lauric acid (31 mg, 0.16
mmol), EDCI (29 mg, 0.16 mmol), and HOBt (21 mg, 0.16 mmol)
in CH2Cl2 (0.8 mL). The reaction mixture was stirred for 12 h.
After removal of the solvent, the resulting residue was directly
purified by column chromatography to give a product (38 mg,
54%) that was then hydrolyzed by NaOMe (0.7 mg, 0.013 mmol)
in MeOH (0.4 mL) and CH2Cl2 (0.1 mL) at room temperature
for 30 min. The mixture was neutralized with IR-120, filtered,
and concentrated. The resulting residue was purified by column
chromatography to give CCL-34-S17 (19.6 mg, 60%). Rf 0.15
N-(2-Acetoxy-2-amino-2-deoxy)-O-(3,4,6-tri-O-acetoxy-b-D-
glucopyranosyl)-N-dodecanoyl-L-serine undecylamide (27)
1
(1 : 15 MeOH–CH2Cl2); H (400 MHz, CDCl3/MeOD, mixture
Acetic acid (1.0 mL, 17.46 mmol) and acetic anhydride (2 mL,
21.20 mmol) were added at room temperature to a solution
of 25 (91.5 mg, 0.10 mmol) and Zn (0.26 g, 3.98 mmol) in
tetrahydrofuran (3.0 ml). The reaction mixture was stirred for
12 h and then filtered over Celite. Tetrahydrofuran, acetic acid,
and acetic anhydride were then removed by reduced pressure.
The resulting residue was purified by column chromatography
to give pure 27 (55.6 mg, 72%). Rf 0.55 (1 : 9 MeOH–CH2Cl2); 1H
(400 MHz, CDCl3/MeOD, mixture of two conformation isomer)
d 0.86 (6H, t, J = 6.7, 7.0 Hz), 1.19–1.36 (32H, m, CH2), 1.47–1.51
(2H, m), 1.57–1.61 (2H, m), 1.89 and 1.90 (3H, s), 1.98 and 1.99
(3H, s), 2.01 (3H, s), 2.06 and 2.07 (3H, s), 2.17–2.30 (2H, m),
3.01–3.28 (2H, m), 3.70– 3.75 (4H, m), 4.13 (1H, dd, J = 2.2, 9.8
Hz), 4.30 (1H, dd, J = 6.1, 12.4 Hz), 4.50 and 4.52 (1H, t, J = 5.8
and 6.1 Hz), 4.63 and 4.67 (1H, d, J = 8.5 and 8.5 Hz), 4.99 and
4.99 (1H, t, J = 9.4 and 9.4 Hz), 5.14 and 5.17 (1H, dd, J = 9.4, 10.5
and 9.4, 10.4 Hz); 13C (100 MHz, CDCl3/MeOD) d 29.92, 30.06,
30.12, 30.26, 30.34, 32.65, 36.78, 40.38, 40.47, 40.60, 48.56, 48.78,
48.99, 49.20, 49.41, 49.84, 53.63, 53.70, 54.49, 54.70, 62.85, 69.56,
69.87, 72.50, 72.61, 3.35, 73.56, 78.08, 78.71, 78.73, 101.98, 102.10,
170.75, 170.84, 171.01, 171.54, 171.89, 171.94, 172.80, 173.17,
175.44, 175.54; HRMS (APCI) calc. for C40H71N3O11Na [M +
Na]+ 792.4986, found 792.4992.
of two conformation isomer) d 0.86 (9H, t, J = 6.7, 7.0 Hz), 1.19–
1.38 (48H, m), 1.48 (2H, bs), 1.60 (4H, bs), 2.18–2.27 (4H, m),
3.06–3.19 (2H, m), 3.22–3.28 (2H, m), 3.36–3.42 (1H, m), 3.59–
3.69 (2H, m), 3.69–3.73 (1H, m), 3.81–3.98 (2H, m), 4.32 and 4.36
(1H, d, J = 8.4 and 8.3 Hz), 4.47–4.51 (1H, m); 13C (100 MHz,
CDCl3/MeOD) d 14.63, 23.72, 26.79, 26.90, 26.98, 27.97, 28.03,
30.26, 30.31, 30.40, 30.45, 30.55, 30.65, 30.67, 30.70, 30.74, 30.76,
30.80, 33.04, 36.98, 37.00, 37.54, 40.67, 40.74, 54.66, 54.74, 56.71,
56.82, 69.52, 69.81, 72.01, 72.17, 75.82, 77.79, 78.68, 79.01, 79.33,
102.55, 103.01, 171.83, 171.97, 176.03, 176.22, 177.06, 177.10;
HRMS (ESI) calc. for C44H85N3O8Na [M + Na]+ 806.6234, found
806.6238.
N-(2-Acetoxy-2-amino-2-deoxy)-O-b-D-glucopyranosyl-N-
dodecanoyl-L-serine undecylamide (CCL-34-S18)
NaOMe (1.2 mg, 0.02 mmol) was added at room temperature
to a solution of 27 (55 mg, 0.07 mmol) in MeOH (0.4 mL) and
CH2Cl2 (0.1 mL). The reaction mixture was stirred for 30 min,
neutralized with IR-120, filtered, and concentrated. The resulting
residue was purified by column chromatography to give CCL-34-
S18 (37.7 mg, 82%). Rf 0.13 (1 : 9 MeOH–CH2Cl2); 1H (400 MHz,
CDCl3/MeOD, mixture of two conformation isomer) d 0.87 (6H,
t, J = 6.7, 7.0 Hz), 1.17–1.45 (32H, m), 1.45–1.49 (2H, m), 1.58–
1.62 (2H, m), 1.97 and 1.99 (3H, s), 2.21–2.25 (2H, m), 3.16 (2H, t,
J = 7.2 Hz), 3.26–3.29 (2H, m), 3.39–3.44 (1H, m), 3.59–3.68 (2H,
m), 3.73 (1H, dd, J = 5.9, 10.3 Hz), 3.83–3.89 (1H, m), 3.94 (1H,
dd, J = 5.2 and 10.3 Hz), 4.31 and 4.37 (1H, d, J = 8.3 and 8.3
Hz), 4.48–4.51 (1H, m); 13C (100 MHz, CDCl3/MeOD) d 14.45,
23.41, 26.48, 27.68, 29.95, 30.11, 30.19, 30.31, 30.39, 32.70, 36.83,
40.45, 54.17, 56.75, 62.53, 69.25, 71.79, 75.58, 77.9, 102.06, 102.60,
171.46, 173.79, 175.67; HRMS (ESI) calc. for C34H65N3O8Na [M +
Na]+ 666.4669, found 806.4674.
N-(2-Amino-2-deoxy)-O-b-D-glucopyranosyl-N-dodecanoyl-L-
serine undecylamide (CCL-34-S16)
NaOMe (3.0 mg, 0.06 mmol) was added at room temperature to
a solution of 26 (105 mg, 0.14 mmol) in MeOH (3.0 mL) and
CH2Cl2 (1.0 mL). The reaction mixture was stirred for 30 min,
neutralized with IR-120, filtered, concentrated, and purified by
column chromatography to give pure CCL-34-S16 (48.6 mg, 60%).
1
Rf 0.38 (1 : 6 MeOH–CH2Cl2); H (400 MHz, CDCl3/MeOD) d
0.88 (3H, t, J = 7.0 Hz), 0.88 (3H, t, J = 7.0 Hz), 1.21–1.40 (32H,
m), 1.46–1.55 (2H, m), 1.56–1.67 (2H, m), 2.26 (2H, t, J = 7.4 Hz),
2.56 (1H, t, J = 8.4 Hz), 3.18 (2H, t, J = 6.4 Hz), 3.21–3.28 (3H,
m), 3.67 (1H, dd, J = 5.3, 11.7 Hz), 3.75 (1H, dd, J = 5.2, 10.3
Hz), 3.88 (1H, d, J = 11.7 Hz), 4.07 (1H, dd, J = 5.5, 10.3 Hz),
4.25 (1H, d, J = 8.4 Hz), 4.56 (1H, t, J = 5.3 Hz); 13C (100 MHz,
CDCl3/MeOD) d 14.60, 23.66, 26.77, 27.90, 30.27, 30.39, 30.47,
30.59, 30.67, 32.97, 36.92, 40.59, 54.70, 54.88, 57.97, 62.55, 70.21,
70.56, 71.53, 71.69, 76.94, 78.00, 103.79, 104.24, 171.70, 176.22;
HRMS (ESI) calc. for C32H64N3O7Na [M + H]+ 602.4744, found
602.4736.
N-Dodecanoyl-L-serine undecylamide (CCL-34-S19)
CCL-34-S19 was obtained from commercial Fmoc-Ser-OH by
coupling with undecylamine, deprotection of the Fmoc group, and
coupling with lauric acid in 66% yield over 3 steps. 1H (400 MHz,
CDCl3) d 0.85 (3H, t, J = 6.7 Hz), 0.85 (3H, t, J = 7.0 Hz), 1.19–
1.31 (H, br s), 1.42–1.49 (2H, m), 1.54–1.65 (2H, m), 2.22 (1H, t,
J = 7.5), 2.23 (1H, t, J = 7.5), 3.19 (1H, td, J = 7.1), 3.20 (1H, td,
J = 5.6, 7.1), 3.56 (1H, dd, J = 4.6, 11.5), 4.09 (1H, dd, J = 3.0, J =
11.5), 4.34–4.39 (1H, ddd, J = 3.0, 4.6, 7.1), 6.69 (1H, d, J = 7.1),
6.92 (1H, t, J = 5.6); 13C (100 MHz, CDCl3) d 14.05, 22.63, 25.59,
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Org. Biomol. Chem., 2011, 9, 2492–2504 | 2503
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26.84, 29.20, 29.30, 29.46, 29.51, 29.57, 31.86, 36.48, 39.53, 53.76,
62.90, 170.89, 174.33. HRMS (ESI) calc. for C26H53N2O3[M +
H]+441.4056, found 441.4051.
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The present work was financially supported by the National
Tsing Hua University, Academia Sinica, and the National Science
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