Synthesis of β-hederin and Hederacolchiside A1: Triterpenoid saponins bearing a unique cytotoxicity-inducing disaccharide moiety

Article abstract of DOI:10.1016/j.carres.2005.10.015

A facile synthetic approach toward oleanolic acid glycoside bearing α-l-rhamnopyranosyl-(1→2)-α-l-arabinopyranosyl moiety, a unique oligosaccharide that strongly induces antitumor activity of oleanane-type triterpenoid saponins, was developed. Based on th

Full text of DOI:10.1016/j.carres.2005.10.015

Carbohydrate
RESEARCH
Carbohydrate Research 341 (2006) 60–67
Synthesis of b-hederin and Hederacolchiside A1: triterpenoid
saponins bearing a unique cytotoxicity-inducing disaccharide moiety
Mao-Sheng Cheng,* Mao-Cai Yan, Yang Liu, Li-Gang Zheng and Jiao Liu
Key Lab of New Drugs Design and Discovery of Liaoning Province, School of Pharmaceutical Engineering,
Shenyang Pharmaceutical University, Shenyang 110016, PR China
Received 1 August 2005; received in revised form 2 October 2005; accepted 30 October 2005
Available online 17 November 2005
Abstract—A facile synthetic approach toward oleanolic acid glycoside bearing a-L-rhamnopyranosyl-(1!2)-a-L-arabinopyranosyl
moiety, a unique oligosaccharide that strongly induces antitumor activity of oleanane-type triterpenoid saponins, was developed.
Based on this approach b-hederin (oleanolic acid 3-O-a-L-rhamnopyranosyl-(1!2)-a-L-arabinopyranoside) was efficiently prepared
from oleanolic acid through stepwise glycosylation in linear eight steps with 52% overall yield, while Hederacolchiside A1 (oleanolic
acid 3-O-a-L-rhamnopyranosyl-(1!2)-[b-D-glucopyranosyl-(1!4)]-a-L-arabinopyranoside) in linear 13 steps with 20% overall yield.
ꢀ 2005 Published by Elsevier Ltd.
Keywords: b-Hederin; Hederacolchiside A1; a-L-Rhamnopyranosyl-(1!2)-a-L-arabinopyranoside; Cytotoxicity-inducing oligosaccharide; Triterpe-
noid saponin; Stepwise glycosylation
1. Introduction
b-Hederin (1) (Fig. 1), namely oleanolic acid 3-O-a-L-
rhamnopyranosyl-(1!2)-a-L-arabinopyranoside, is
a
oleanane-type triterpenoid saponin widely distributed
in the nature.1–3 The most attractive property of b-hed-
erin is its prominent cytotoxicity to various human
tumor cell lines such as HL-60 (IC50 = 4.4 lg/mL) and
A549, PC3, DLD1, M4 Beu, PA 1, etc.4,5 Notably,
many oleanane-type triterpenoid saponins bearing a-L-
rhamnopyranosyl-(1!2)-a-L-arabinopyranosyl moiety
at 3-OH (for instance, 2–7) generally have significant
antitumor activity.4–7 For a distinguished example,
Hederacolchiside A (hederagenin 3-O-a-L-rhamnopyr-
anosyl-(1!2)-[b-D-glucopyranosyl-(1!4)]-a-L-arabino-
pyranoside, 2) has been used for chemotherapy of solid
tumor.8 Hederacolchiside A1 (oleanolic acid 3-O-a-L-
rhamnopyranosyl-(1!2)-[b-D-glucopyranosyl-(1!4)]-
a-L-arabinopyranoside, 3)2,9 has been found to have bet-
ter activity than 1 and 2 to all seven tumor cell lines
investigated in the literature.5 Cytotoxicity of their agly-
Figure 1. Glcp- = b-D-glucopyranosyl; Xylp- = b-D-xylopyranosyl.
cons, oleanolic acid (8) and hederagenin (9) (Fig. 2), is
very weak, and their glycosides without a-L-rhamnopyr-
anosyl-(1!2)-a-L-arabinopyranosyl disaccharide moi-
ety at C-3 commonly show much weaker activity.
*
Corresponding author. Tel.: +86 24 23986413; fax: +86 24
23995043; e-mail: mscheng@syphu.edu.cn
0008-6215/$ - see front matter ꢀ 2005 Published by Elsevier Ltd.
doi:10.1016/j.carres.2005.10.015

M.-S. Cheng et al. / Carbohydrate Research 341 (2006) 60–67
61
due to the steric hinderence of sugar acceptor.15,16
Therefore, newly evolved protecting group AZMB(2-
azidomethylbenzoyl)17,18 is often employed in (1!2)-
linkage construction,19,20 for it is stable enough to pre-
vent the by-product forming and can be selectively re-
moved under the treatment of Bu3P. However,
selective introduction of a special neighboring partici-
pating protective group such as AZMB to C-2 of sugar
donors is rather boring in preparation of numerous
glycosides with various saccharide moiety. A linear syn-
thetic route employing the readily available perbenzoy-
lated sugar donors is more efficient and convenient for
this work.
Accordingly, perbenzoylated trichloroacetimidate
1121 and 1522 (Scheme 1) were chosen as sugar donors,
which can be easily prepared from L-arabinose and L-
rhamnose, respectively. Ester linkage at C-28 of olean-
olic acid is resistant to basic hydrolysis due to its high
steric hinderence, making it difficult and inefficient to
deprotect after the glycosylation.23 Trityl,21 tert-butyldi-
phenylsilyl,19 and allyl24 have been employed to protect
the carboxyl of oleanolic acid in previous work. Here,
we used benzyl ester (10) instead which can be prepared
under the treatment of BnBr and Et3N in dry THF at a
high yield and easily removed through catalytic hydro-
genation (double bond between C-12 and C-13 of olean-
olic acid is inert to catalytic hydrogenation25).
As is shown in Scheme 1, esterification of oleanolic
acid (8) followed by glycosylation with trichloroacetim-
idate 11 under the promotion of TMSOTf gave 12 in an
excellent yield. Debenzoylation of 12 in NaOMe/MeOH
without influencing benzyl ester at C-28 produced 13.
Selective shelter of hydroxyl groups at C-3 and C-4 of
arabinose residue was successfully carried out using
2,2-dimethoxypropane.26 Coupling of 14 and 15 pro-
moted by TMSOTf led to partly breakage of isopropy-
lidene group, so glycosylation of 14 was carried out
under the promotion of BF3ÆEt2O at a low temperature
in 79% yield. We had hoped that isopropylidene could
be removed automatically after being warmed to rt,
but the products became complicated and could not be
separated very well through silica gel column chroma-
tography. So Et3N was added after glycosylation to neu-
tralize the Lewis acid and an additional step of
deprotection was performed to provide intermediate
17. Finally, ready removal of benzyl group through cat-
alytic hydrogenation and benzoyl group in NaOMe/
MeOH afforded 1, whose physical data are in well agree-
ment with the literature reported.1–3
Figure 2.
Therefore, the a-L-rhamnopyranosyl-(1!2)-a-L-arabi-
nopyranosyl disaccharide moiety has proved to be a un-
ique sugar sequence which strongly induces antitumor
activity of oleanane-type glycosides4–7 and has been
attracting more and more attention of researchers.
Several hypotheses have been suggested for the mech-
anism of their antitumor activity.10–12 Initial structure–
activity relationship (SAR) study has shown that a
glucopyranosyl moiety attached to C-4 of arabinose in-
creases the cytotoxicity against malignant melanoma
M4 Beu markedly,5 while methylation of C-28 carboxyl
of the aglycon does not affect the cytotoxicity against
HL-60 significantly.4 However, research in this domain
is sporadic and unsystematic mainly due to the difficulty
in obtaining adequate analogical glycosides from nature
for broad and thorough SAR study. Based on our expe-
rience with synthesis and bioactivity study of antitumor
natural products,13 we get excited about preparing b-
hederin and a sufficiency of analogues and derivatives
through chemical synthesis for SAR investigation and
development of desirable antitumor agents.
2. Results and discussion
So far no synthetic approach toward oleanolic acid gly-
coside bearing this unique disaccharide, a-L-rhamnopyr-
anosyl-(1!2)-a-L-arabinopyranosyl moiety, has been
reported yet. This disaccharide bears a (1!2)-linkage,
thus the usual strategy of preparing the disaccharide do-
nor followed by coupling with the aglycon is not adopt-
able, otherwise a mixture of a- and b-anomers would be
afforded without a neighboring participating group on
C-2 of arabinose residue. Alternately, stepwise glycosyl-
ation generally affords the exclusive 1,2-trans-glycoside
linkage so long as a neighboring participating group is
on C-2 of sugar donor. Moreover, this strategy provides
a versatile approach for preparing a glycoside family
with variety simply by altering monosaccharide do-
nors.14 Therefore, stepwise glycosylation was adopted
in our work.
Hederacolchiside A1 (3) was further synthesized from
key intermediate 17 (Scheme 2). Selective protection of
hydroxyl group on equatorial bond of 1,2-diol may be
achieved by various methods such as BzCl27 and Bu2S-
nO/BzCl.28 It is known that cyclic ortho ester formed
from 1,2-diol and CH3C(OEt)3 cleaving under acidic
condition generally results in monoacetylation of axial
It is known that glycosylation of C-3 of triterpenoid
requires a benzoyl at C-2 of sugar donor, otherwise a
low yield would be obtained with the ortho ester forma-
tion and the acyl transfer being serious side reactions

62
M.-S. Cheng et al. / Carbohydrate Research 341 (2006) 60–67
˚
Scheme 1. Reagents and conditions: (a) BnBr, Et3N, Bu4NI, THF, rt, 97%; (b) TMSOTf, CH2Cl2, MS 4 A, rt, 94%; (c) NaOMe, CH2Cl2–MeOH, rt,
˚
97%; (d) Me2C(OMe)2, TsOH, acetone, 0 ꢁC to rt, 89%; (e) BF3ÆEt2O, CH2Cl2, MS 4 A, ꢀ78 ꢁC, 79%; (f) TsOH, CH2Cl2–MeOH, rt, 98%; (g) Pd–C,
H2, EtOAc, reflux; (h) NaOMe, CH2Cl2–MeOH, rt, 86% (for two steps).
Scheme 2. Reagents and conditions: (a) CH3C(OEt)3, TsOH, toluene, rt; (b) 80% aq HOAc, rt, 89% (for two steps); (c) BzCl, pyridine, 0 ꢁC to rt; (d)
˚
AcCl, CH2Cl2–MeOH, 0 ꢁC to rt, 74% (for two steps); (e) TMSOTf, CH2Cl2, MS 4 A, rt, 71%; (f) Pd–C, H2, EtOAc, reflux; (g) NaOMe, CH2Cl2–
MeOH, rt, 70% (for two steps).
hydroxyl group in excellent yield.29,30 Herein, diol 17
was converted into cyclic ortho ester 19 and then cleaved
in aq HOAc to furnish 20, which would be useful in
preparation of advanced derivatives glycosylated at C-
3 of arabinose residue (for instance, 7) in future work.
Benzoylation of 20 and selective deacetylation at the
presence of benzoyl group31 gave 22. Free hydroxyl of
22 was then glycosylated by 2,3,4,6-tetra-O-benzoyl-a-
D-glucopyranosyl trichloroacetimidate 2332 under the
promotion of TMSOTf. Excessive 23 (2.50 equiv) and
promoter (0.40 equiv) were needed in this step to pro-
vide a satisfactory yield (71%). The final removal of ben-
zyl and benzoyl furnished 3, whose physical data are
identical with those reported for the natural product.2,9
In conclusion, a facile synthetic approach toward
oleanolic acid glycoside bearing a-L-rhamnopyranosyl-
(1!2)-a-L-arabinopyranosyl moiety at C-3 was devel-
oped employing perbenzoylated trichloroacetimidates

M.-S. Cheng et al. / Carbohydrate Research 341 (2006) 60–67
63
of monosaccharide as sugar donors. b-Hederin was pre-
3.3. Benzyl oleanolate 3-O-2,3,4-tri-O-benzoyl-a-L-
arabinopyranoside (12)
pared from oleanolic acid in linear eight steps at an
overall yield of 52%. Hederacolchiside A1 was further
synthesized from intermediate 17 through seven steps
at a yield of 33% (20% overall yield from oleanolic acid).
More advanced monodesmosides and bisdesmosides can
be readily synthesized from intermediates 17, 18, 20, 22,
and 25. This approach is competent for high efficient
preparation of analogues and derivatives on a large scale
for structure–activity relationship investigation and new
chemical entity finding. Further study on preparation
and bioactivity evaluation of analogues and derivatives
is currently underway.
Benzyl ester 10 (895 mg, 1.64 mmol), trichloroacetimi-
date 11 (1140 mg, 1.88 mmol) and powdered 4 A
˚
molecular sieves (1600 mg) were stirred for 40 min at
rt in dry CH2Cl2 (18 mL). A dry CH2Cl2 solution
(1.60 mL) of TMSOTf (0.016 mL, 0.082 mmol) was
added dropwise. The mixture was stirred for 20 min
followed by addition of Et3N (0.20 mL) and filtration.
The filtrate was concentrated and purified by a silica
gel column chromatography (8:1, petroleum ether–
EtOAc) to afford 12 (1620 mg, 94%) as a white foam.
[a]D +108.2 (c 1.63, CHCl3); Rf = 0.65 (4:1, petroleum
ether–EtOAc); 1H NMR (300 MHz, CDCl3): d 8.09–
7.26 (m, 20H, Ar–H), 5.75 (dd, J 8.8, 6.5, 1H, H-20),
5.67 (m, 1H, H-40), 5.59 (dd, J 8.9, 3.5, 1H, H-30),
5.30 (t, J 3.0, 1H, H-12), 5.06 (dd, J 19.9, 12.6, 2H,
PhCH2), 4.79 (d, J 6.6, 1H, H-10), 4.33 (dd, J 12.9,
3.8, 1H, H-50-1), 3.87 (m, 1H, H-50-2), 3.16 (dd, J
11.0, 4.9, 1H, H-3), 2.90 (dd, J 10.1, 3.2, 1H, H-18),
1.11, 0.92, 0.90, 0.86, 0.79, 0.66, 0.58 (s each, 3H each,
7 · Me); 13C NMR (75 MHz, CDCl3): d 177.4, 165.8,
165.6, 165.2, 143.6, 136.4, 133.3, 133.2, 133.1, 129.9,
129.8, 129.7, 129.4, 129.1, 128.4, 128.4, 128.3, 127.9,
127.8, 122.5, 103.1, 90.1, 70.7, 70.2, 68.6, 65.9, 62.6,
55.4, 47.5, 46.7, 45.8, 41.6, 41.3, 39.2, 38.9, 38.4,
36.7, 33.8, 33.1, 32.6, 32.3, 30.7, 27.8, 27.6, 25.8,
23.6, 23.3, 23.0, 18.1, 16.8, 16.2, 15.2; ESI-MS (m/z):
1013.5 (M+Na).
3. Experimental
3.1. General methods
Commercial reagents were used without further purifica-
tion unless specialized. Solvents were dried and redis-
tilled prior to use in the usual way. Boiling range of
petroleum ether was 60–90 ꢁC. Analytical TLC was per-
formed with silica gel HF254. Preparation column chro-
matography was performed with silica gel H. Melting
¨
points were detected with BUCHI Melting Point
B-540. Optical rotations were measured at the sodium
D-line at room temperature with a Perkin–Elmer 241
MC polarimeter. 1H and 13C NMR spectra were
recorded on Bruker ARX 300 MHz or Avance AV
600 MHz, using Me4Si as the internal standard if not
specially mentioned. J Values are given in hertz. ESI-
MS were obtained on an Agilent 1100 mass spectrome-
ter. High-resolution mass spectra (FT-ICRMS) were de-
tected on Bruker APEX II mass spectrometer.
3.4. Benzyl oleanolate 3-O-a-L-arabinopyranoside (13)
To a solution of 12 (1100 mg, 1.11 mmol) in dry
CH2Cl2–MeOH (1:2, 40 mL) was added a newly pre-
pared NaOMe in MeOH solution (1.0 mol/L,
1.20 mL). The mixture was stirred at rt for 2 h and neu-
tralized with Dowex H+ resin to pH 7 and then filtered.
The filtrate was concentrated and the resulting residue
was subjected to a silica gel column chromatography
(EtOAc) to gave 13 (734 mg, 97%) as a white amorphous
solid. [a]D +37.5 (c 0.77, acetone); Rf = 0.35 (EtOAc);
1H NMR (300 MHz, Me2SO-d6): d 7.33 (m, 5H,
PhCH2), 5.17 (br s, 1H, H-12), 5.02 (m, 2H, PhCH2),
4.80 (br s, 1H, OH), 4.53 (br s, 1H, OH), 4.47 (br s,
1H, OH), 4.09 (d, J 6.0, 1H, H-10), 3.60 (m, 2H, H-50-
1, H-40), 3.35–3.32 (m, 3H, H-20, H-30, H-50-2), 2.99
(dd, J 10.5, 3.2, 1H, H-3), 2.80 (dd, J 9.8, 3.0, 1H, H-
18), 1.07, 0.94, 0.87, 0.85, 0.82, 0.73, 0.51 (s each, 3H
each, 7 · Me); 13C NMR (75 MHz, Me2SO-d6): d
176.4, 143.4, 136.4, 128.5, 128.0, 127.8, 122.1, 106.0,
87.7, 72.8, 71.1, 67.8, 65.4, 65.3, 55.0, 47.0, 46.2, 45.5,
41.3, 41.1, 38.1, 36.3, 33.2, 32.8, 32.4, 32.1, 30.5, 27.7,
27.1, 25.8, 25.6, 23.4, 23.0, 22.7, 17.8, 16.7, 16.5, 15.2;
ESI-MS (m/z): 701.4 (M+Na).
3.2. Benzyl oleanolate (10)
A solution of oleanolic acid (1070 mg, 2.34 mmol), BnBr
(0.39 mL, 3.28 mmol), Et3N (0.45 mL, 3.28 mmol) and
Bu4NI (86 mg, 0.24 mmol) in dry THF (20 mL) was stir-
red at rt overnight. The solvent was evaporated in vac-
uum and the residue was purified through a silica gel
column chromatography (8:1, petroleum ether–EtOAc)
to give 10 (1240 mg, 97%) as a white amorphous solid.
[a]D +50.1 (c 2.44, CHCl3); Rf = 0.55 (4:1, petroleum
ether–EtOAc); 1H NMR (300 MHz, CDCl3): d 7.33
(m, 5H, PhCH2), 5.29 (t, J 3.0, 1H, H-12), 5.07 (dd, J
17.8, 12.6, 2H, PhCH2), 3.21 (dd, J 10.3, 4.8, 1H, H-
3), 2.91 (dd, J 14.0, 4.2, 1H, H-18), 1.13, 0.99, 0.92,
0.90, 0.88, 0.78, 0.61 (s each, 3H each, 7 · Me); 13C
NMR (75 MHz, CDCl3): d 177.5, 143.7, 136.4, 128.4,
128.0, 127.9, 122.5, 79.0, 65.9, 55.2, 47.6, 46.7, 45.9,
41.7, 41.4, 39.2, 38.7, 38.4, 37.0, 33.9, 33.1, 32.7, 32.4,
30.7, 28.1, 27.6, 27.2, 25.9, 23.6, 23.4, 23.0, 18.3, 16.9,
15.6, 15.3; ESI-MS (m/z): 1115.9 (2M+Na).

64
M.-S. Cheng et al. / Carbohydrate Research 341 (2006) 60–67
3.5. Benzyl oleanolate 3-O-3,4-O-isopropylidene-a-L-
arabinopyranoside (14)
129.7, 129.6, 129.5, 129.4, 129.3, 128.5, 128.4, 128.3,
128.2, 127.9, 122.5, 110.4, 103.3, 95.2, 89.1, 79.2, 75.3,
73.4, 72.0, 70.6, 69.9, 66.5, 65.9, 62.7, 55.8, 47.6, 46.7,
45.8, 41.7, 41.4, 39.3, 39.2, 38.7, 36.7, 33.8, 33.1, 32.7,
32.4, 30.7, 28.2, 27.8, 27.6, 26.1, 25.9, 23.6, 23.4, 23.0,
18.1, 17.5, 16.9, 16.7, 15.3; ESI-MS (m/z): 1200.0
(M+Na); HRMS: calcd for C65H81O14 (MꢀBn):
1085.5626; found: m/z 1085.5619.
To a solution of compound 13 (679 mg, 1.00 mmol) in
dry acetone (12 mL) stirred at 0 ꢁC was added Me2-
C(OMe)2 (0.31 mL, 2.50 mmol) and TsOH (17.2 mg).
The mixture was then allowed to warm up to rt and stir-
red for 4 h before Et3N (0.20 mL) was added. The solu-
tion was concentrated and purified through a silica gel
column chromatography (6:1, petroleum ether–EtOAc)
to afford 14 (634 mg, 89%) as a white foam. [a]D +45.0
(c 1.60, CHCl3); Rf = 0.45 (4:1, petroleum ether–
3.7. Benzyl oleanolate 3-O-2,3,4-tri-O-benzoyl-a-L-
rhamnopyranosyl-(1!2)-a-L-arabinopyranoside (17)
1
EtOAc); H NMR (300 MHz, CDCl3): d 7.34 (m, 5H,
TsOH (78 mg) was added to a solution of 16 (705 mg,
0.600 mmol) in CH2Cl2–MeOH (1:2, 45 mL) and the
solution was stirred at rt. When TLC (2:1, petroleum
ether–EtOAc) showed that deprotection had completed,
Et3N (0.40 mL) was added and the mixture was concen-
trated and purified through a silica gel column chroma-
tography (2:1, petroleum ether–EtOAc) to give 17
(666 mg, 98%) as a white amorphous solid. [a]D +77.3
(c 2.27, CHCl3); Rf = 0.36 (2:1, petroleum ether–
PhCH2), 5.28 (t, J 3.0, 1H, H-12), 5.07 (dd, J 18.7,
12.6, 2H, PhCH2), 4.22–4.17 (m, 3H, H-10, H-40, H-50-
1), 4.06 (dd, J 7.8, 6.1, 1H, H-30), 3.75 (dd, J 13.9, 3.5,
1H, H-50-2), 3.63 (dd, J 7.8, 7.8, 1H, H-20), 3.12 (dd, J
11.5, 4.6, 1H, H-3), 2.91 (dd, J 13.8, 3.3, 1H, H-18),
2.30 (br s, 1H, OH), 1.54, 1.36 (s each, 3H each, O–
(CH3)2C–O), 1.11, 0.98, 0.92, 0.89, 0.88, 0.82, 0.60 (s
each, 3H each, 7 · Me); 13C NMR (75 MHz, CDCl3):
d 177.4, 143.6, 136.3, 128.3, 127.9, 127.8, 122.4, 110.0,
104.3, 88.9, 78.1, 74.2, 73.2, 65.9, 63.0, 55.4, 47.5, 46.6,
45.8, 41.6, 41.3, 39.2, 39.0, 38.4, 36.6, 33.8, 33.1, 32.6,
32.3, 30.6, 28.2, 28.0, 27.5, 26.0, 25.8, 23.6, 23.3, 22.9,
18.1, 16.8, 16.6, 15.2; ESI-MS (m/z): 741.4 (M+Na);
HRMS: calcd for C38H59O7 (MꢀBn): 627.4261; found:
m/z 627.4255.
1
EtOAc); H NMR (300 MHz, CDCl3): d 8.10–7.23 (m,
20H, Ar–H), 5.84 (dd, J 10.2, 3.1, 1H, H-300), 5.65 (m,
2H, H-200, H-400), 5.36 (s, 1H, H-100), 5.29 (br s, 1H, H-
12), 5.07 (dd, J 18.7, 12.6, 2H, PhCH2), 4.81 (d, J 1.3,
1H, H-10), 4.34 (m, 1H, H-500), 4.11–3.98 (m, 3H, H-20,
H-40, OH), 3.82 (m, 1H, H-50-1), 3.67 (m, 1H, H-50-2),
3.45 (d, J 7.9, 1H, H-30), 3.18 (dd, J 11.0, 3.2, 1H, H-
3), 2.91 (m, 1H, H-18), 2.52 (br s, 1H, OH), 1.34 (d, J
6.1, 3H, H-600), 1.12, 1.05, 0.92, 0.89, 0.88, 0.84, 0.61 (s
each, 3H each, 7 · Me); 13C NMR (75 MHz, CDCl3):
d 177.4, 165.7, 165.5, 143.6, 136.4, 133.5, 133.3, 133.1,
129.9, 129.7, 129.6, 129.2, 129.2, 129.1, 128.5, 128.4,
128.2, 127.9, 127.8, 122.4, 102.0, 98.2, 90.3, 76.2, 71.5,
70.7, 70.7, 69.7, 67.3, 65.9, 65.5, 61.1, 55.4, 47.6, 46.7,
45.8, 41.6, 41.3, 39.2, 39.1, 38.5, 36.7, 33.8, 33.1, 32.6,
32.3, 30.6, 28.1, 27.6, 25.8, 25.7, 23.6, 23.4, 23.0, 18.2,
17.5, 16.8, 16.4, 15.3; ESI-MS (m/z): 1159.6 (M+Na).
3.6. Benzyl oleanolate 3-O-2,3,4-tri-O-benzoyl-a-L-
rhamnopyranosyl-(1!2)-3,4-O-isopropylidene-a-L-
arabinopyranoside (16)
A suspension of 14 (560 mg, 0.779 mmol), 15 (630 mg,
˚
1.01 mmol) and powdered 4 A molecular sieves
(850 mg) in dry CH2Cl2 (8 mL) was stirred for 40 min
and then cooled to ꢀ78 ꢁC. BF3ÆEt2O (0.070 mL,
0.545 mmol) was added and the mixture was stirred at
ꢀ78 ꢁC for 2 h before the reaction was quenched by
Et3N (0.20 mL). The suspension was then filtered and
the filtrate was concentrated and subjected to a silica
gel chromatography (8:1, petroleum ether–EtOAc) to
furnish 16 (730 mg, 79%) as a white foam. [a]D +96.7
(c 2.58, CHCl3); Rf = 0.60 (4:1, petroleum ether–
3.8. Oleanolic acid 3-O-a-L-rhamnopyranosyl-(1!2)-
a-L-arabinopyranoside (1)
A suspension of 17 (63 mg, 0.055 mmol) and 10% Pd–C
(30 mg) in EtOAc (16 mL) was refluxed and bubbled up
with H2 (20 mL/min) for 3 h. The mixture was then
filtered and the filtrate was concentrated to dryness to
afford crude 18 as a colorless oil. Crude 18 was dissolved
in dry CH2Cl2–MeOH (1:2, 18 mL), to which a newly
prepared NaOMe in MeOH solution (1.0 mol/L,
0.80 mL) was added. The mixture was stirred at rt for
2 h and then neutralized with Dowex H+ resin to pH 7
and filtered. The filtrate was concentrated and purified
1
EtOAc); H NMR (300 MHz, CDCl3): d 8.12–7.21 (m,
20H, Ar–H), 5.87 (dd, J 10.2, 3.3, 1H, H-300), 5.76 (br
s, 1H, H-100), 5.65 (m, 2H, H-200, H-400), 5.30 (t, J 3.0,
1H, H-12), 5.07 (dd, J 22.4, 12.6, 2H, PhCH2), 4.53
(m, 1H, H-500), 4.47 (d, J 3.0, 1H, H-10), 4.25 (m, 2H,
H-30, H-40), 4.17 (m, 1H, H-50-1), 3.90 (dd, J 3.0, 3.0,
1H, H-20), 3.79 (m, 1H, H-50-2), 3.17 (dd, J 11.3, 4.1,
1H, H-3), 2.92 (m, 1H, H-18), 1.55, 1.35 (s each, 3H
each, O–(CH3)2C–O), 1.34 (d, J 6.1, 3H, H-600), 1.14,
0.95, 0.93, 0.92, 0.90, 0.89, 0.64 (s each, 3H each,
7 · Me); 13C NMR (75 MHz, CDCl3): d 177.4, 165.7,
165.5, 165.4, 143.7, 136.4, 133.3, 133.2, 132.9, 130.0,
with
a silica gel column chromatography (15:1,
EtOAc–MeOH) to gave 1 (35 mg, 86%) as a white pow-
der. Mp 237–240 ꢁC, lit.1 222–225 ꢁC, lit.2 230–240 ꢁC,
lit.3 240–241 ꢁC; [a]D +4.5 (c 0.60, MeOH), lit.1 +9 (c

M.-S. Cheng et al. / Carbohydrate Research 341 (2006) 60–67
65
1.83, MeOH), lit.2 +6 (c 1.0, MeOH), lit.3 +4.29 (c 0.35,
MeOH); Rf = 0.23 (7:1, CHCl3–MeOH); 1H NMR
(300 MHz, pyridine-d5, solvent peak as internal stan-
dard (7.57 ppm)): d 6.15 (s, 1H, H-100), 5.46 (br s, 1H,
H-12), 4.88 (d, J 5.0, 1H, H-10), 4.74 (m, 1H, H-200),
4.64–4.56 (m, 3H, H-20, H-300, H-500), 4.33–4.27 (m,
4H, H-30, H-40, H-50-1, H-400), 3.82 (m, 1H, H-50-2),
3.26 (m, 2H, H-3, H-18), 1.61 (d, J 5.7, 3H, H-600),
1.29, 1.17, 1.06, 1.00, 0.98, 0.95, 0.82 (s each, 3H each);
13C NMR (150 MHz, pyridine-d5, solvent peak as inter-
nal standard (135.6 ppm)): d 180.3 (C-28), 144.9 (C-13),
122.6 (C-12), 105.0 (C-10), 101.8 (C-100), 88.8 (C-3), 76.0
(C-20), 74.1 (C-400), 74.0 (C-30), 72.6 (C-300), 72.5 (C-200),
69.9 (C-500), 68.8 (C-40), 64.9 (C-50), 56.0 (C-5), 48.1 (C-
9), 46.7 (C-17), 46.5 (C-19), 42.2 (C-14), 42.0 (C-18),
39.8 (C-4), 39.6 (C-8), 38.9 (C-1), 37.1 (C-10), 34.3 (C-
21), 33.4, 33.3, 33.2 (C-7, C-22, C-29), 31.0 (C-20),
28.4 (C-15), 28.1 (C-23), 26.6 (C-2), 26.2 (C-27), 23.8,
23.7, 23.7 (C-11, C-16, C-30), 18.6, 18.6 (C-6, C-600),
17.4 (C-26), 17.1 (C-24), 15.6 (C-25); ESI-MS (m/z):
757.5 (M+Na); HRMS: calcd for C41H65O11 (MꢀH):
733.4527; found: m/z 733.4529.
27.6, 25.8, 23.6, 23.4, 23.0, 21.1, 18.2, 17.5, 16.8, 16.5,
15.3; ESI-MS (m/z): 1201.7 (M+Na); HRMS: calcd for
C64H79O15 (MꢀBn): 1087.5419; found: m/z 1087.5419.
3.10. Benzyl oleanolate 3-O-2,3,4-tri-O-benzoyl-a-L-
rhamnopyranosyl-(1!2)-3-O-benzoyl-a-L-arabinopyr-
anoside (22)
A solution of 20 (240 mg, 0.203 mmol) in dry pyridine
(5 mL) was stirred and cooled to 0 ꢁC. After slow addi-
tion of BzCl (0.070 mL, 0.610 mmol), the mixture was
warmed up to rt and stirred overnight. Water
(0.40 mL) was added slowly to quench the reaction
and the solvent was evaporated in vacuum. The result-
ing residue was dissolved in CH2Cl2 (20 mL) and
washed with water (15 mL · 3) and dried over MgSO4.
The mixture was filtered and the filtrate was concen-
trated to afford crude 21 as a white amorphous solid,
which was then dissolved in dry CH2Cl2–MeOH (1:2,
15 mL). To the solution was added AcCl (0.30 mL) at
0 ꢁC, and the solution was warmed to rt and stirred until
TLC (3:1, petroleum ether–EtOAc) showed the starting
material had disappeared. Et3N (1.20 mL) was added to
neutralize the acid. The solution was then concentrated
and purified with a silica gel column chromatography
(5:1, petroleum ether–EtOAc) to furnish 22 (187 mg,
74%) as a white foam. [a]D +88.7 (c 2.32, CHCl3);
Rf = 0.40 (3:1, petroleum ether–EtOAc); 1H NMR
(300 MHz, CDCl3): d 8.08–7.19 (m, 25H, Ar–H), 5.80
(dd, J 10.2, 3.3, 1H, H-300), 5.64 (m, 2H, H-200, H-400),
5.36 (s, 1H, H-100), 5.34–5.29 (m, 2H, H-30, H-12), 5.07
(dd, J 20.0, 12.6, 2H, PhCH2), 4.76 (d, J 4.1, 1H, H-
10), 4.41 (m, 1H, H-500), 4.33–4.27 (m, 2H, H-20, H-40),
4.08 (m, 1H, H-50-1), 3.72 (dd, J 10.5, 2.7, 1H, H-50-
2), 3.17 (dd, J 11.5, 3.8, 1H, H-3), 2.91 (dd, J 10.3,
3.6, 1H, H-18), 1.33 (d, J 6.1, 3H, H-600), 1.12, 1.03,
0.92, 0.90, 0.87, 0.70, 0.60 (s each, 3H each, 7 · Me);
13C NMR (150 MHz, CDCl3): d 177.4, 165.9, 165.7,
165.2, 165.2, 143.7, 136.4, 133.5, 133.3, 133.3, 133.2,
133.0, 130.1, 129.9, 129.8, 129.7, 129.6, 129.5, 129.5,
129.2, 129.1, 128.5, 128.4, 128.4, 128.2, 128.1, 128.0,
127.9, 122.5, 102.7, 98.1, 89.3, 73.8, 73.8, 71.7, 70.6,
69.5, 67.3, 65.9, 65.5, 62.6, 55.6, 47.6, 46.7, 45.8, 41.6,
41.3, 39.3, 39.2, 38.7, 36.7, 33.8, 33.1, 32.6, 32.3, 30.7,
29.7, 28.0, 27.6, 26.0, 25.8, 23.6, 23.4, 23.0, 18.1, 17.4,
16.8, 16.4, 15.4; ESI-MS (m/z): 1263.7 (M+Na); HRMS:
calcd for C69H81O15 (MꢀBn): 1149.5576; found: m/z
1149.5569.
3.9. Benzyl oleanolate 3-O-2,3,4-tri-O-benzoyl-a-L-
rhamnopyranosyl-(1!2)-4-O-acetyl-a-L-arabino-
pyranoside (20)
A solution of 17 (285 mg, 0.250 mmol), CH3C(OEt)3
(0.23 mL, 1.25 mmol) and TsOH (16 mg) in dry toluene
(8 mL) was stirred at rt for 1 h before the reaction was
quenched with Et3N (0.2 mL). The mixture was diluted
with toluene (20 mL) and washed with water
(20 mL · 3). The solvent was removed in vacuum to fur-
nish crude 19 as a colorless oil, which was then dissolved
in 80% aq HOAc (8 mL) and stirred at rt for another
1 h. Coevaporation with toluene yielded a straw yellow
solid, which was then subjected to a silica gel column
chromatography (5:1, petroleum ether–EtOAc) to give
20 (262 mg, 89%) as a white foam. [a]D +85.8 (c 2.28,
CHCl3); Rf = 0.27 (3:1, petroleum ether–EtOAc); 1H
NMR (300 MHz, CDCl3): d 8.10–7.22 (m, 20H, Ar–
H), 5.86 (m, 1H, H-300), 5.75–5.65 (m, 2H, H-200, H-400),
5.47 (s, 1H, H-100), 5.29 (t, J 2.5, 1H, H-12), 5.14 (m,
1H, H-40), 5.10 (dd, J 20.1, 12.6, 2H, PhCH2), 4.72 (d,
J 4.5, 1H, H-10), 4.43 (m, 1H, H-500), 4.12 (m, 1H, H-
30), 3.98 (m, 1H, H-50-1), 3.95 (m, 1H, H-20), 3.69 (dd,
J 12.1, 3.0, 1H, H-50-2), 3.35 (br s, 1H, OH), 3.19 (dd,
J 11.3, 4.4, 1H, H-3), 2.91 (dd, J 12.3, 2.8, 1H, H-18),
2.15 (s, 3H, CH3CO), 1.35 (d, J 6.3, 3H, H-600), 1.12,
1.10, 0.92, 0.89, 0.89, 0.88, 0.61 (s each, 3H each,
7 · Me); 13C NMR (150 MHz, CDCl3): d 177.4, 170.8,
165.8, 165.5, 165.5, 143.7, 136.4, 133.4, 133.3, 133.1,
129.9, 129.8, 129.7, 128.5, 128.4, 128.2, 127.9, 127.9,
122.4, 103.0, 98.2, 90.3, 76.2, 71.6, 70.7, 70.5, 69.8,
69.6, 67.2, 65.9, 59.9, 55.6, 47.6, 46.7, 45.8, 41.6, 41.3,
39.3, 39.1, 38.6, 36.7, 33.8, 33.1, 32.6, 32.3, 30.7, 28.1,
3.11. Benzyl oleanolate 3-O-2,3,4-tri-O-benzoyl-a-L-
rhamnopyranosyl-(1!2)-[2,3,4,6-tetra-O-benzoyl-b-D-
glucopyranosyl-(1!4)]-3-O-benzoyl-a-L-arabinopyrano-
side (24)
A suspension of 22 (160 mg, 0.129 mmol), 23 (240 mg,
˚
0.322 mmol) and powdered 4 A molecular sieves

66
M.-S. Cheng et al. / Carbohydrate Research 341 (2006) 60–67
(400 mg) in dry CH2Cl2 (6 mL) were stirred for 40 min
at rt. A dry CH2Cl2 solution (1.00 mL) of TMSOTf
(0.010 mL, 0.052 mmol) was added dropwise. The mix-
ture was stirred for 1 h before Et3N (0.16 mL) was
added to quench the reaction. The mixture was then di-
luted with CH2Cl2 (25 mL) and filtered. The filtrate was
concentrated and purified by a silica gel column chroma-
tography (5:1, petroleum ether–EtOAc) to afford 24
(166 mg, 71%) as a white foam. [a]D +47.0 (c 1.38,
CHCl3); Rf = 0.42 (4:1, petroleum ether–EtOAc); 1H
NMR (600 MHz, CDCl3): d 8.08–7.10 (m, 45H, Ar–
H), 5.88 (dd, J 9.6, 9.6, 1H, H-3000), 5.78 (m, 1H, H-
300), 5.73 (dd, J 9.6, 9.6, H-4000), 5.64 (m, 2H, H-200, H-
400), 5.55 (m, 1H, H-2000), 5.27 (m, 1H, H-12), 5.21–5.05
(m, 4H, PhCH2, H-100, H-1000), 4.71 (m, 2H, H-10, H-
30), 4.53 (dd, J 9.0, 1.4, 1H, H-20), 4.43 (m, 1H, H-6000-
1), 4.33 (m, 2H, H-40, H-500), 4.25–4.20 (m, 3H, H-50-1,
H-5000, H-6000-2), 3.77 (m, 1H, H-50-2), 3.06 (m, 1H, H-
3), 2.83 (dd, J 13.3, 3.1, 1H, H-18), 1.33 (d, J 6.3, 3H,
H-600), 1.10, 1.04, 0.93, 0.90, 0.87, 0.81, 0.57 (s each,
3H each, 7 · Me); 13C NMR (75 MHz, CDCl3): d
177.4, 166.0, 165.8, 165.7, 165.6, 165.4, 165.2, 165.1,
165.0, 143.6, 136.4, 133.3, 133.0, 132.8, 129.9, 129.8,
129.2, 129.1, 128.9, 128.8, 128.4, 128.2, 128.1, 127.9,
122.5, 101.8, 100.6, 98.2, 89.2, 73.7, 73.7, 72.8, 72.1,
71.6, 71.6, 70.5, 70.3, 69.7, 69.6, 67.3, 65.9, 62.8, 60.4,
55.5, 47.6, 46.7, 45.8, 41.6, 41.3, 39.3, 39.1, 38.6, 36.7,
33.8, 33.1, 32.6, 32.4, 30.7, 28.0, 27.8, 27.6, 25.8, 23.6,
23.4, 23.0, 18.1, 17.5, 16.8, 16.2, 15.3; ESI-MS (m/z):
50-1), 4.30–4.16 (m, 4H, H-30, H-40, H-3000, H-4000), 4.03
(dd, J 7.9, 7.9, 1H, H-2000), 3.91 (m, 1H, H-5000), 3.80 (d,
J 11.3, 1H, H-50-2), 3.27–3.22 (m, 2H, H-3, H-18),
1.64 (d, J 6.2, 3H, H-600), 1.28, 1.17, 1.10, 1.00, 0.97,
0.94, 0.82 (s each, 3H each, 7 · Me); 13C NMR
(150 MHz, pyridine-d5, solvent peak as internal stan-
dard (135.6 ppm)): d 180.2 (C-28), 144.9 (C-13), 122.6
(C-12), 106.4 (C-1000), 105.0 (C-10), 101.8 (C-100), 88.7
(C-3), 79.7 (C-40), 78.8 (C-5000), 78.6 (C-3000), 76.4 (C-20),
75.5 (C-2000), 74.1, 74.1 (C-30, C-400), 72.5, 72.4 (C-200,
C-300), 71.3 (C-4000), 69.9 (C-500), 64.6 (C-50), 62.6 (C-
6000), 56.0 (C-5), 48.1 (C-9), 46.7 (C-17), 46.5 (C-19),
42.2 (C-14), 42.0 (C-18), 39.8 (C-4), 39.5 (C-8), 38.9
(C-1), 37.1 (C-10), 34.3 (C-21), 33.3, 33.2, 33.2 (C-7,
C-22, C-29), 31.0 (C-20), 28.4 (C-15), 28.1 (C-23), 26.6
(C-2), 26.2 (C-27), 23.8, 23.7, 23.7 (C-11, C-16, C-30),
18.7 (C-600), 18.5 (C-6), 17.4 (C-26), 17.1 (C-24),
15.6 (C-25); ESI-MS (m/z): 919.5 (M+Na); HRMS:
calcd for C47H75O16 (MꢀH): 895.5055; found: m/z
895.5050.
References
1. Higuchi, R.; Kawasaki, T. Chem. Pharm. Bull. 1976, 24,
1021–1032.
2. Schenkel, E. P.; Werner, W.; Schulte, K. E. Planta Med.
1991, 57, 463–467.
3. Zhong, H. M.; Chen, C. X.; Tian, X.; Chui, Y. X.; Chen,
Y. Z. Planta Med. 2001, 67, 484–488.
1842.7 (M+Na); HRMS: calcd for
(MꢀBn): 1727.7152; found: m/z 1727.7149.
C103H107O24
4. Mimaki, Y.; Kuroda, M.; Asano, T.; Sashida, Y. J. Nat.
Prod. 1999, 62, 1279–1283.
5. Barthomeuf, C.; Debiton, E.; Mshvidadze, V.; Kemerte-
lidze, E.; Balansard, G. Planta Med. 2002, 68, 672–675.
6. Jung, H. J.; Lee, C. O.; Lee, K. T.; Choi, J.; Park, H. J.
Biol. Pharm. Bull. 2004, 27, 744–747.
7. Park, H. J.; Kwon, S. H.; Lee, J. H.; Lee, K. H.;
Miyamoto, K.; Lee, K. T. Planta Med. 2001, 67, 118–121.
8. Kim, S. B.; Ahn, B. Z.; Kim, Y. Eur. Patent 1,388,542,
2004; Chem. Abstr. 2004, 140, 139489u.
9. Ekabo, O. A.; Farnsworth, N. R. J. Nat. Prod. 1996, 59,
431–435.
3.12. Oleanolic acid 3-O-a-L-rhamnopyranosyl-(1!2)-
[b-D-glucopyranosyl-(1!4)]-a-L-arabinopyranoside (3)
A suspension of 24 (122 mg, 0.067 mmol) and 10% Pd–C
(45 mg) in EtOAc (18 mL) was refluxed and bubbled up
with H2 (20 mL/min) for 4 h. Pd–C was removed
through filtration and the filtrate was concentrated to
dryness. The resulting residue was dissolved in dry
CH2Cl2–MeOH (1:2, 24 mL), to which a newly prepared
NaOMe in MeOH solution (1.0 mol/L, 1.00 mL) was
added. The solution was stirred at rt for 2 h and then
neutralized with Dowex H+ resin to pH 7 and filtered.
The filtrate was concentrated and subjected to a silica
gel column chromatography (20:10:1, CHCl3–MeOH–
H2O) to give a pale yellow powder, which was recrystal-
lized in MeOH to afford 3 (42 mg, 70%) as a white pow-
der. Mp 251–253 ꢁC, lit.2 250–260 ꢁC; [a]D ꢀ2.1 (c 0.16,
MeOH), lit.2 0 (c 2.97, MeOH), lit.9 ꢀ6.11 (c 0.135,
EtOH); Rf = 0.35 (20:10:1, CHCl3–MeOH–H2O); 1H
NMR (300 MHz, pyridine-d5, solvent peak as internal
standard (7.56 ppm)): d 6.20 (s, 1H, H-100), 5.46 (m,
1H, H-12), 5.14 (d, J 7.8, 1H, H-1000), 4.77–4.73 (m,
2H, H-10, H-200), 4.67–4.60 (m, 3H, H-20, H-300, H-500),
4.53–4.48 (m, 2H, H-6000), 4.41–4.34 (m, 2H, H-400, H-
10. Jeong, H. G.; Choi, C. Y. Planta Med. 2002, 68, 392–
396.
11. Barthomeuf, C.; Boivin, D. Cancer Chemother. Pharma-
col. 2004, 54, 432–440.
12. Debiton, E.; Borel, M.; Communal, Y.; Mshvildadze, V.;
Barthomeuf, C. Melanoma Res. 2004, 14, 97–105.
13. Cheng, M. S.; Wang, Q. L.; Tian, Q.; Song, H. Y.; Liu, Y.
X.; Li, Q.; Xu, X.; Miao, H. D.; Yao, X. S.; Yang, Z. J.
Org. Chem. 2003, 68, 3658–3662.
14. Yu, B.; Hui, Y. In Glycochemistry: Principles, Synthesis
and Applications; Wang, P. G., Bertozzi, C. R., Eds.;
Marcel Dekker: New York, 2001; pp 167–174.
15. Urban, F. J.; Moore, B. S.; Breitenbach, R. Tetrahedron
Lett. 1990, 31, 4421–4424.
16. Deng, S.; Yu, B.; Xie, J.; Hui, Y. J. Org. Chem. 1999, 64,
7265–7266.
17. Wada, T.; Ohkubo, A.; Mochizuki, A.; Sekine, M.
Tetrahedron Lett. 2001, 42, 1069–1072.
18. Love, K. R.; Andrade, R. B.; Seeberger, P. H. J. Org.
Chem. 2001, 66, 8165–8176.

M.-S. Cheng et al. / Carbohydrate Research 341 (2006) 60–67
67
19. Peng, W.; Sun, J.; Lin, F.; Han, X.; Yu, B. Synlett 2004,
259–262.
26. Nicolaou, K. C.; Trujillo, J. I.; Chibale, K. Tetrahedron
1997, 53, 8751–8778.
20. Peng, W.; Han, X.; Yu, B. Synthesis 2004, 1641–1647.
21. Yu, B.; Xie, J.; Deng, S.; Hui, Y. J. Am. Chem. Soc. 1999,
121, 12196–12197.
22. Ziegler, T.; Bien, F.; Jurisch, C. Tetrahedron: Asymmetry
1998, 9, 765–780.
27. Kozikowski, A. P.; Yan, X.; Rusnak, J. M. J. Chem. Soc.,
Chem. Commun. 1988, 1301–1303.
28. David, S.; Hanessian, S. Tetrahedron 1985, 41, 643–663.
29. Hanessian, S.; Roy, R. Can. J. Chem. 1985, 63, 163–
172.
23. Ohtani, K.; Mizutani, K.; Kasai, R.; Tanaka, O. Tetra-
hedron Lett. 1984, 25, 4537–4540.
30. Bouchra, M.; Calinaud, P.; Gelas, J. Carbohydr. Res.
1995, 267, 227–237.
24. Sun, J.; Han, X.; Yu, B. Carbohydr. Res. 2003, 338, 827–
833.
25. Winterstein, A.; Stein, G. Z. Physiol. Chem. 1931, 202,
222–231.
31. Byramova, N. E.; Ovchinnikov, M. V.; Bakinovskii, L. V.;
Kochetkov, N. K. Carbohydr. Res. 1983, 124, C8–C11.
32. Schmidt, R. R.; Kinzy, W. Adv. Carbohydr. Chem.
Biochem. 1994, 50, 21–125.