Efficient access to isomeric 2,3-dihydroxy lupanes: first synthesis of alphitolic acid

Article abstract of DOI:10.1016/j.tet.2009.07.047

Alphitolic acid (3) is a naturally occurring lupane type of pentacyclic triterpene, which possesses various pharmacological properties. Efficient synthesis of 3 has been accomplished in 10 steps with an overall yield of 19% starting from the readily avail

Full text of DOI:10.1016/j.tet.2009.07.047

Tetrahedron 65 (2009) 7975–7984
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Efficient access to isomeric 2,3-dihydroxy lupanes: first synthesis of alphitolic acid
,
Jia Hao a, Xueli Zhang a, Pu Zhang a, Jun Liu b, Luyong Zhang b, Hongbin Sun a
*
a Center for Drug Discovery, College of Pharmacy, China Pharmaceutical University, 24 Tongjia Xiang, Nanjing 210009, China
b Jiangsu Center for Drug Screening, China Pharmaceutical University, 1 Shennonglu, Nanjing 210038, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 28 May 2009
Received in revised form 17 July 2009
Accepted 22 July 2009
Available online 25 July 2009
Alphitolic acid (3) is a naturally occurring lupane type of pentacyclic triterpene, which possesses various
pharmacological properties. Efficient synthesis of 3 has been accomplished in 10 steps with an overall
yield of 19% starting from the readily available diketone 11. An alternative approach to the key in-
termediate 17 has also been developed, and based on this approach, 3 could be obtained in 10 steps with
an overall yield of 26% starting from 11. Moreover, seven other isomeric 2,3-dihydroxy lupanes 4–10 have
been synthesized. The synthesized triterpenes 3–10 were evaluated for their inhibitory activity against
rabbit muscle GPa (RMGPa), and some of them exhibited moderate inhibitory activity against RMGPa.
Ó 2009 Elsevier Ltd. All rights reserved.
1. Introduction
1
2
OH
OH
R1= H
R1=
R2=
R2=
OH R2=
OH R2=
OH R2=
X=H2
X=O
29
H
20
30
OH
OH
3 R1=
X=O
Betulin (1) and betulinic acid (2) (Fig. 1), two well-known
members of lupane type of pentacyclic triterpenes, have recently
attracted much attention due to their anti-cancer and anti-HIV
activities.1 An ointment containing 2 is currently under phase II
clinical trial for treatment of dysplastic nevi.2 Previously, we
reported that 1, 2 and other naturally occurring pentacyclic tri-
terpenes, represented a novel class of inhibitors of glycogen phos-
19
17
15
4 R1=
5 R1=
6 R1=
7 R1=
8 R1=
9 R1=
X=H2
H
11
OH X=O
OH
25
2613
1
4
28
OH
OH
OH
OH
OH
R2=
X=H2
X=O
X=H2
X=O
R1
R2
X
2
3
10
8
OH R2=
OH R2=
OH R2=
27
24
23
OH
10 R1=
R2=
OH X=H2
phorylases (GP).3 Alphitolic acid (2
a-hydroxybetulinic acid, 3),
which possesses various pharmacological properties such as anti-
bacterial, anti-inflammatory, antiproliferative, antiplasmodial, anti-
HIV activities,4–8 was firstly isolated from the ethereal extract of
Alphitonia whitei Braid.9,10 In this study, we developed efficient
synthetic approaches to 3 and related isomeric 2,3-dihydroxy
lupanes 4–1011,12 so as to assist further pharmaceutical research on
these biologically important molecules. To the best of our knowl-
edge, this is the first synthesis of 3. Moreover, the synthesized tri-
terpenes 3–10 were evaluated for their inhibitory activity against
rabbit muscle GPa (RMGPa).
Figure 1. Alphitolic acid (3) and related triterpenes.
m-CPBA catalyzed by H2SO4, or by hydroboration–oxidation of the
corresponding enol acetates.13,14 Treatment of lupanes under the
similar conditions, however, would affect the iso-propylene group
to afford 2,30-dihydroxy compounds.15 When using Pb(AcO)4 as an
oxidant to build up the 2a-hydroxy function in betulin derivatives,
the iso-propylene group was still affected to result in 2,30-diac-
etyloxy products.16 The high reactivity of the iso-propylene func-
tion group in lupanes thereby seemingly made it a difficult task to
selectively carry out hydroxylation at C-2. Herein, we report new
access to isomeric 2,3-dihydroxy lupanes 3–10.
The synthetic approach to 3 is shown in Scheme 1. Treatment of
diketone 11, which was readily prepared from 1,17 with tert-butyl-
dimethylsilylchloride (TBDMSCl) and imidazole in DMF gave enol
silyl ether 12 in quantitative yield. Reduction of 12 with sodium
2. Results and discussion
2.1. Chemistry
In previous studies, introduction of the 2a-hydroxy function
into oleananes and ursanes could be accomplished by stereo-
selective hydroxylation of the corresponding 3-ketones with
borohydride afforded 2
b
-hydroxy-3b-O-silylated product 13 as the
major product (80%), together with the 2
b
-O-silylated product 14 as
a minor product (14%). Obviously, a transsilylation occurred during
this reduction reaction. The migration of the TBS group from the
C-2 oxygen atom to its C-3 neighbor might have taken place via an
enediolate intermediate.18 The relative configurations of 13 and 14
* Corresponding author. Tel.: þ86 25 85327950; fax: þ86 25 83271335.
E-mail address: hbsun2000@yahoo.com (H. Sun).
0040-4020/$ – see front matter Ó 2009 Elsevier Ltd. All rights reserved.
doi:10.1016/j.tet.2009.07.047

7976
J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
the minor product (38%). Removal of the TBS group in 16 with
tetrabutylammonium fluoride (TBAF) in refluxing THF gave 2 ,3b-
a
H
H
diol 17 (94%). Acetylation of 17 with acetic anhydride in the pres-
ence of 4-dimethylaminopyridine (DMAP) in pyridine, followed by
deprotection with PPTS afforded alcohol 18 in 76% yield for two
steps. Oxidation of 18 with pyridinium chlorochromate (PCC) gave
aldehyde 19 (96%), which was further oxidized with sodium chlo-
rite (NaClO2) and sodium dihydrogenophosphate in a mixture of
t-BuOH/THF/2-methyl-2-butene20 to furnish triterpene acid 20
(82%). Hydrolysis of 20 with aqueous sodium hydroxide in meth-
anol gave alphitolic acid (3) in 97% yield. Deprotection of 17 with
CH2OH
lit. 17
CH2OTr
TBSCl,
imidazole, DMF
45 oC, 99%
HO
O
HO
1 (betulin)
11
H
H
CH2OTr
CH2OTr
NaBH4,
R1O
R2O
TBSO
O
EtOH, THF
rt
PPTS afforded 2
yield.
a,3b,28-lup-20(29)-en-triol (4) in quantitative
12
13 R1=H, R2=TBS, major (80%)
14 R1=TBS, R2=H, minor (14%)
On the other hand, an alternative approach to the key in-
termediate 17 has also been developed. As shown in Scheme 2,
Meerwein–Pondorf reduction of 11 gave 2
product (60%), together with 17 as the minor product (22%).
Deprotection of 21 with PPTS afforded 2 ,3 ,28-lup-20(29)-en-triol
(6) in 85% yield. Selective protection of 21 with tert-butyldime-
thylsilylchloride (TBDMSCl) and imidazole in DMF gave 2-TBS ether
a,3a-diol 21 as the major
H
Al(i-PrO)3, i-PrOH,
CH2OTr
for 13
PCC, CH2Cl2
0 oC, 78%
O
AlCl3 (cat.)
a
a
reflux, 57
%
TBSO
15
22 in high yield (92%) due to a less steric hindrance at 2
than that at 3 -hydroxy. Oxidation of 22 with pyridinium chloro-
chromate (PCC) at 0 ꢀC followed by sodium borohydride reduction
afforded 3 -alcohol 24 (81%, for two steps). Deprotection of 24 with
tetrabutylammonium fluoride (TBAF) in refluxing THF gave 17
(99%).
a-hydroxy
a
H
H
b
CH2OTr
CH2OTr
HO
TBSO
HO
HO
TBAF, THF
reflux, 94%
16
17
H
H
1) Ac2O, pyridine, DMAP
2) PPTS, EtOH, 70 o
TBSCl,
imidazole, DMF
Al(i-PrO)3, i-PrOH,
CH2OH
CH2OTr
AcO
HO
HO
C
PCC, CH2Cl2
rt, 96%
AlCl3 (cat.)
11
reflux, 60%
rt, 92%
76% for two steps
AcO
18
21
6
PPTS, EtOH
70 oC, 85%
H
H
H
H
NaH2PO4, NaClO2
t-BuOH, THF,
2-methyl-2-butene
CH2OTr
CH2OTr
PCC, CH2Cl2
0 o
CO2H
CHO
TBSO
TBSO
HO
AcO
AcO
AcO
82%
C
O
AcO
20
19
22
23
H
H
NaBH4,
EtOH, THF
CH2OTr
CO2H
TBSO
HO
HO
HO
TBAF, THF
reflux, 99%
NaOH (aq.), MeOH
rt, 97%
17
rt
81% from 22
3
24
Scheme 2. Synthesis 2a,3a,28-lup-20(29)-en-triol (6) and an alternative approach to
the key intermediate 17.
H
CH2OH
HO
HO
PPTS, EtOH
70 oC, 99%.
In the above regards, two synthetic approaches to 3 have been
established starting from 11. For the first approach, which
employed a reduction–transsilylation as the key step (Scheme 1), 3
was obtained in 10 steps with an overall yield of 19%. Based on the
second approach, which started with stereoselective Meerwein–
Pondorf reduction of 11 (Scheme 2), 3 was gained in 10 steps with
a total yield of 26%.
17
4
Scheme 1. Synthesis of alphitolic acid (3) and 2a,3b,28-lup-20(29)-en-triol (4).
have been confirmed by NOE experiments (see Supplementary
data). It’s interesting that 2 -hydroxy-3 -O-silylated product 13
could be obtained in high regioselectivity and stereoselectivity. Our
previous studies showed that C2 –OH was likely less steric hin-
dered than C3 ,3 -diols
–OH21, and therefore, direct silylation of 2
could provide 2 -hydroxy-3 -O-silylated products only as the mi-
nor products, together with 2
b
b
2b,3a,28-Lup-20(29)-en-triol (8) was prepared according to the
methodology developed for preparation of bredemolic acid
(Scheme 3).21 Therefore, treatment of diol 2517 with tert-butyldi-
methylsilylchloride (TBDMSCl) and imidazole in DMF gave the
2-silyl ether 14 as the major product (82%), together with the 3-silyl
ether 13 as a minor product (11%). Oxidation of 14 with pyridinium
chlorochromate (PCC) at 0 ꢀC followed by Meerwein–Pondorf re-
duction afforded alcohol 27 (41% for two steps). Deprotection of 27
with tetrabutylammonium fluoride (TBAF) in refluxing THF gave
b
b
b b
b
b
b
-O-silylated products as the major
products (>70%).21 Oxidation of 13 with pyridinium chloro-
chromate (PCC) at 0 ꢀC produced 2-ketone 15 in 78% yield. Meer-
wein–Pondorf reduction19 of 15 afforded 2
a
-hydroxy isomer 16 as
the major product (57%), together with the 2
b-hydroxy isomer 13 as
2b,3a-diol 28 (90%). Deprotection of 28 with PPTS afforded

J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
7977
2
b
,3
a
-dihydroxy-lup-20(29)-en-28-ol (8) (97%). On the other hand,
35, 36, and 37 with aqueous sodium hydroxide in methanol gave 5
(96%), 9 (96%), and 7 (95%), respectively.
deprotection of 25 with PPTS afforded
20(29)-en-28-ol (10) (98%).
2b
,3b
-dihydroxy-lup-
2.2. Biological activity
The synthesized 2,3-dihydroxy lupanes 3–10 were evaluated for
their inhibitory activity against rabbit muscle GPa (RMGPa). The
activity of RMGPa was measured by detecting the release of
phosphate from glucose-1-phosphate in the direction of glycogen
synthesis.22 The bioassay results (Table 1) showed that most of the
2,3-dihydroxy lupanes exhibited moderate inhibition against
H
H
TBSCl,
imidazole, DMF
40 oC, 82%
CH2OTr
CH2OTr
HO
HO
TBSO
HO
14
25
10
PPTS, EtOH
70 oC, 98%
RMGPa. Naturally occurring triterpenes 3 (IC50 20.7
mM) and 5 (IC50
H
13.1 M) were more potent inhibitors than the other synthetic
triterpenes. Further biological evaluation of these triterpenes is
warranted to determine their therapeutic potential.
m
Al(i-PrO)3, i-PrOH,
AlCl3 (cat.)
CH2OTr
PCC, CH2Cl2 TBSO
oC, 78%
reflux, 53%
0
O
26
Table 1
IC50 values (
mM) for the inhibition of rabbit muscle GPa
H
H
a
Compound
GPa IC50
CH2OTr
CH2OTr
HO
HO
TBSO
HO
1
2
3
41.5
20.9
20.7
98.5
13.1
499
26.9
105.5
28.4
34.2
98.5
TBAF, THF
reflux, 90%
27
4
5
6
7
8
9
PPTS, EtOH
70 oC, 97%
28
8
Scheme 3. Synthesis of 2
20(29)-en-28-ol (10).
b,3a,28-lup-20(29)-en-triol (8) and 2b,3b-dihydroxy-lup-
Using the same methodology for the conversion of 17 to 3
(Scheme 1), isomeric 2,3-dihydroxy lupane acids 5, 7, and 9 were
synthesized starting from 21, 28, and 25, respectively (Scheme 4).
Acetylation of 21, 25, and 28 with acetic anhydride in the presence
of 4-dimethylaminopyridine (DMAP) in anhydrous pyridine, fol-
lowed by deprotection with PPTS afforded alcohol 29 (73%), 30
(72%), and 31 (71%), respectively. Oxidation of 29, 30, and 31 with
pyridinium chlorochromate (PCC) gave aldehyde 32 (87%), 33
(89%), and 34 (92%), respectively. Oxidation of 32, 33, and 34 with
sodium chlorite (NaClO2) and sodium dihydrogenophosphate in
a mixture of t-BuOH/THF/2-methyl-2-butene furnished triterpene
acids 35 (84%), 36 (86%), and 37 (88%), respectively. Hydrolysis of
10
Caffeine
a
Values are means of three experiments.
3. Conclusions
In summary, efficient synthesis of alphitolic acid (3) has been
accomplished in 10 steps with an overall yield of 19% starting from
the readily available diketone 11. An alternative approach to the key
intermediate 17 has also been developed, and based on this ap-
proach, 3 could be obtained in 10 steps with an overall yield of 26%
starting from the same material. Seven other isomeric 2,3-dihy-
droxy lupanes 4–10 have also been synthesized. The synthesized
triterpenes have been biologically evaluated as inhibitors of gly-
cogen phosphorylase, and the result shows that naturally occurring
H
H
triterpenes 3 (IC50 20.7 mM) and 5 (IC50 13.1 mM) are moderate GP
inhibitors.
CH2OTr
CH2OH
R1
R2
R1
R2
1) Ac2O, pyridine
DMAP
2) PPTS, EtOH
70 oC
4. Experimental section
4.1. General
OH
21 R1=
25 R1=
28 R1=
OH, R2=
29 R1=
30 R1=
OAc,
OAc,
OAc,
OAc (73%)
OAc (72%)
R2=
OH,
OH
OH
R2=
R2=
R =
2
OH,
31 R1=
OAc (71%)
R2=
H
All commercially available solvents and reagents were used
without further purification. Melting points are uncorrected. 1H
NMR and 13C NMR spectra were recorded at 300 and 75 MHz, re-
spectively. Chemical shifts are reported as values from an internal
tetramethylsilane standard. Low- and high-resolution mass spectra
(LRMS and HRMS) were given with electron impact mode. Infrared
spectra were recorded either on neat samples (KBr disks) or as thin
film.
CHO
NaH2PO4, NaClO2
PCC, CH2Cl2
rt
R1
R2
t-BuOH, THF,
2-methyl-2-butene
OAc,
OAc, R2=
OAc, R2=
32 R1=
33 R1=
34 R1=
R2=
OAc (87%)
OAc (89%)
OAc (92%)
H
H
4.2. 28-Trityl-2-(tert-butyldimethylsilyloxy)-3-oxo-lup-
1,20(29)-dien-28-ol (12)
CO2H
CO2H
R1
R2
R1
NaOH (aq.), MeOH
rt
R2
Compound 1117 (1 g, 1.44 mmol) was dissolved in 10 mL of DMF,
then imidazole (801 mg, 11.78 mmol) and tert-butyl dimethysilyl
chloride (887 mg, 5.89 mmol) were added. The reaction mixture
was stirred at 45 ꢀC for 2 h. The mixture was cooled to rt, then
10 mL of ice water was added to the reaction mixture, the mixture
OH,
OH,
OH,
OH (96%)
OH (96%)
OH (95%)
5 R1=
9 R1=
7 R1=
R2=
R2=
R2=
OAc (84%)
OAc (86%)
OAc (88%)
35 R1=
36 R1=
37 R1=
OAc, R2=
OAc,
OAc,
R2=
R2=
Scheme 4. Synthesis of isomeric 2,3-dihydroxy lupane acids 5, 7, and 9.

7978
J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
was extracted with ethyl acetate (4ꢁ50 mL). The combined extract
was washed with brine, dried over anhydrous sodium sulfate,
filtered, and the solvent was removed under reduced pressure.
The crude product was purified by column chromatography (pe-
troleum ether–ethyl acetate, 5:1) to give 12 as a white solid (1.15 g,
99%). Mp 247–248 ꢀC; IR (KBr, cmꢂ1) 3068, 2949, 2859, 2352, 1681,
1673, 1620, 1453, 1381, 1254, 1216, 1061, 842, 782, 705, 632, 584; 1H
4.5. 28-Trityl-3
lup-20(29)-en-28-ol (16)
b-(tert-butyldimethylsilyloxy)-2a-hydroxy-
To a solution of 15 (660 mg, 0.812 mmol) in dry isopropanol was
added 12 mL of fresh prepared aluminum isopropoxide
(1.54 mmol/ml) in dry isopropanol and cat. amount of aluminum
chloride, the reaction mixture was stirred at 110 ꢀC for 5 days. The
reaction mixture was cooled to rt, the solvent was removed in
vacuo, and the pH was adjusted to 3.0 by 1 N HCl at 0 ꢀC. The
mixture was extracted with ethyl acetate (4ꢁ40 mL). The combined
extract was washed with saturated sodium bicarbonate solution
and brine, dried over anhydrous sodium sulfate, filtered and the
solvent was removed under reduced pressure. The crude product
was purified by column chromatography (petroleum ether/ethyl
NMR (C6D6N)
d 0.27 (s, 3H), 0.30 (s, 3H), 0.73 (s, 3H), 0.87 (s, 3H),
1.02 (s, 9H), 1.06 (s, 9H), 1.15 (s, 3H), 1.20 (s, 3H), 1.70 (s, 3H), 2.33–
2.43 (m, 3H), 3.17 and 3.39 (d, J¼8.9 Hz, each 1H), 4.70 and 4.76 (br
s, each 1H), 6.61 (s, 1H), 7.30–7.34 (m, 3H), 7.39–7.44 (m, 6H), 7.70–
7.73 (m, 6H); 13C NMR (C6D6N)
d
ꢂ4.3, ꢂ4.1, 14.8, 16.6, 18.8, 19.3,
19.4, 20.6, 21.3, 21.9, 25.6, 26.0, 27.3, 28.3, 30.4, 30.5, 34.0, 35.6, 37.7,
39.2, 41.8, 43.0, 45.4, 45.6, 47.95, 48.02, 49.1, 53.7, 60.0, 86.5, 110.1,
127.4, 128.3, 129.3, 138.0, 145.1, 145.7, 150.8, 200.9; ESI-MS m/z:
811.4 [MþH]þ; HRMS calcd for C55H75O3Si [MþH]þ: 811.5485,
found: 811.5480.
acetate, 120:1) to give 28-trityl-3b-(tert-butyldimethylsilyloxy)-2a-
hydroxy-lup-20(29)-en-28-ol (16) as a white solid (376 mg, 57%).
Mp 101–103 ꢀC; IR (film, cmꢂ1) 2948, 2858, 1448, 1257, 1109, 1088,
1064, 834, 774, 742, 705, 631; 1H NMR (CDCl3)
d 0.04 (s, 3H), 0.05 (s,
4.3. 28-Trityl-3
lup-20(29)-en-28-ol (13)
b
-(tert-butyldimethylsilyloxy)-2
b
-hydroxy-
3H), 0.44 (s, 3H), 0.69 (s, 3H), 0.76 (s, 3H), 0.82 (s, 3H), 0.85 (s, 3H),
0.86 (s, 9H), 1.56 (s, 3H), 1.84–1.89 (m, 1H), 2.10–2.13 (m, 4H), 2.84
and 3.06 (d, J¼8.8 Hz, each 1H), 2.91 (d, J¼9.1 Hz,1H), 3.50–3.55 (m,
1H), 4.46 (br s, 1H), 4.51 (d, J¼1.6 Hz, 1H), 7.12–7.43 (m, 15H); 13C
Compound 12 (240 mg, 0.30 mmol) was dissolved in THF
(5 mL) and EtOH (1 mL), then NaBH4 (13.23 mg, 0.35 mmol) was
added, and the reaction mixture was stirred for 5 h at rt. The
reaction was quenched by 1 N HCl at 0 ꢀC, then organic solvent
was removed under reduced pressure, the mixture was extracted
with ethyl acetate (4ꢁ40 mL). The organic layers were combined
and washed with brine, dried over anhydrous sodium sulfate,
filtered, and the solvent was removed under reduced pressure, the
residue was purified by column chromatography (petroleum
NMR (CDCl3)
d
ꢂ3.2, ꢂ4.3,14.7,16.0,17.0,17.3,18.61, 18.65,19.1, 20.9,
25.1, 26.2, 26.9, 29.0, 30.0, 30.2, 34.2, 35.2, 37.3, 38.1, 40.2, 40.7, 42.5,
46.5, 47.6, 47.8, 49.0, 50.3, 55.4, 59.6, 69.2, 85.6, 85.9, 109.4, 126.8,
127.7, 128.8, 144.5, 150.7; ESI-MS m/z: 813.9 [MꢂH]ꢂ; HRMS calcd
for C55H78NaO3Si [MþNa]þ: 837.5618, found: 837.5612.
4.6. 28-Trityl-2
a,3b-dihydroxy-lup-20(29)-en-28-ol (17)
(prepared from 16)
ether/ethyl acetate, 160:1) to give 28-trityl-3
b
-(tert-butyldime-
white
thylsilyloxy)-2
b
-hydroxy-lup-20(29)-en-28-ol (13) as
a
Compound 16 (180 mg, 0.22 mmol) was dissolved in THF
(12 mL) and two drops of water were added, then 0.56 mL of TBAF
solution (1 M) was added to the solution, the reaction mixture was
heated to reflux, and maintained for 2 h, then the solvent was re-
moved directly, and the residue was purified directly by column
chromatography (petroleum ether/ethyl acetate, 5:1) to give 28-
solid (193 mg, 80%). Mp 134–136 ꢀC; IR (film, cmꢂ1) 3571, 2949,
2854, 1640, 1489, 1470, 1448, 1381, 1252, 1108, 1087, 1062, 877,
837, 744, 765, 705, 631; 1H NMR (CDCl3)
d 0.07 (s, 3H), 0.10 (s, 3H),
0.52 (s, 3H), 0.88 (s, 3H), 0.89 (s, 3H), 0.93 (s, 9H), 0.95 (s, 3H), 1.09
(s, 3H), 1.63 (s, 3H), 2.12–2.24 (m, 5H), 2.48 (br s, 1H), 2.90 and
3.13 (d, J¼8.8 Hz, each 1H), 3.21 (d, J¼3.9 Hz, 1H), 3.88–3.89 (m,
1H), 4.53 and 4.59 (d, J¼2.1 Hz, each 1H), 7.22–7.33 (m, 9H), 7.47–
trityl-2a,3b-dihydroxy-lup-20(29)-en-28-ol (17) as a white solid
(145 mg, 94%). Mp 142–144 ꢀC; IR (film, cmꢂ1) 3400, 3060, 2942,
2867, 1641, 1594, 1489, 1448, 1374, 1264, 1064, 1032, 983, 880, 774,
7.50 (m, 6H); 13C NMR (CDCl3)
d 14.7, 15.9, 17.1, 17.6, 18.2, 19.2,
20.9, 25.3, 25.9, 26.9, 29.7, 29.8, 30.0, 30.2, 34.2, 35.2, 36.6, 37.3,
38.6, 40.7, 42.6, 43.3, 47.6, 47.7, 49.0, 50.9, 55.3, 59.6, 71.6, 80.1,
85.9, 109.3, 126.8, 127.7, 128.8, 144.5, 150.8; ESI-MS m/z: 813.4
[MꢂH]ꢂ; HRMS calcd for C55H78NaO3Si [MþNa]þ: 837.5618,
found: 837.5612.
764, 742, 705, 631; 1H NMR (CDCl3)
d 0.50 (s, 3H), 0.79 (s, 3H), 0.83
(s, 3H), 0.89 (s, 3H), 1.00 (s, 3H),1.63 (s, 3H),1.95–2.00 (m, 1H), 2.12–
2.28 (m, 3H), 2.91 and 3.12 (d, J¼9.0 Hz, each 1H), 2.95 (d, J¼9.5 Hz,
1H), 3.60–3.68 (m, 1H), 4.52 and 4.58 (d, J¼2.1 Hz, each 1H), 7.22–
7.50 (m, 15H); 13C NMR (CDCl3)
d 14.7, 15.9, 16.5, 17.3, 18.3, 19.1, 20.8,
25.0, 26.9, 28.4, 29.9, 30.1, 34.0, 35.2, 37.2, 38.5, 39.2, 40.7, 42.5, 46.7,
47.6, 47.8, 48.9, 50.2, 55.3, 59.5, 69.2, 83.9, 85.8, 109.4, 126.8, 127.7,
128.8, 144.5, 150.7; ESI-MS m/z: 699.5 [MꢂH]ꢂ; HRMS calcd for
C49H64NaO3 [MþNa]þ: 723.4753, found: 723.4747.
4.4. 28-Trityl-3
28-ol-2-one (15)
b-(tert-butyldimethylsilyloxy)-lup-20(29)-en-
To a solution of 13 (280 mg, 0.34 mmol) in dichloromethane was
added PCC (88.36 mg, 0.41 mmol), the reaction mixture was stirred
at 0 ꢀC for 10 h, the mixture was filtered and concentrated in vacuo,
the residue was purified directly by column chromatography (pe-
4.7. 2a,3b-Diacetyloxy-lup-20(29)-en-28-ol (18)
To a solution of 17 (570 mg, 0.81 mmol) in dry pyridine (3 mL)
was added Ac2O (1 mL), the reaction mixture was stirred at rt
overnight. After cooling to 0 ꢀC, 1 N HCl (25 mL) was added into the
reaction mixture. The mixture was extracted with ethyl acetate
(3ꢁ30 mL). The combined extract was washed with saturated so-
dium bicarbonate solution and brine, dried over anhydrous sodium
sulfate, filtered, and concentrated in vacuo. The crude product was
used directly without further purification. The crude residue was
dissolved in absolute ethanol (10 mL) and was then added PPTS
(732 mg, 2.92 mmol). The reaction mixture was stirred at 70 ꢀC for
8 h. After cooling to room temperature, the solvent was evaporated
in vacuo. Ice water (15 mL) was added to the residue, and extracted
with ethyl acetate (3ꢁ20 mL). The combined extract was washed
with brine, dried over anhydrous sodium sulfate, filtered, and
troleum ether/ethyl acetate, 160:1) to give 28-trityl-3b-(tert-
butyldimethylsilyloxy)-lup-20(29)-en-28-ol-2-one (15) as a white
solid (217.8 mg, 78%). Mp 126–128 ꢀC; IR (film, cmꢂ1) 2947, 1724,
1452, 1253, 1152, 1062, 835, 772, 704, 631; 1H NMR (CDCl3)
d
ꢂ0.00
(s, 3H), 0.11 (s, 3H), 0.53 (s, 3H), 0.74 (s, 3H), 0.77 (s, 3H), 0.95 (s, 9H),
0.96 (s, 3H), 1.13 (s, 3H), 1.67 (s, 3H), 2.22–2.36 (m, 4H), 2.95 and
3.15 (d, J¼8.8 Hz, each 1H), 3.94 (s, 1H), 4.56 and 4.62 (br s, each
1H), 7.23–7.53 (m, 15H); 13C NMR (CDCl3)
d
ꢂ5.5, -4.2, 14.7, 15.6,
16.6, 16.7, 19.0, 19.2, 21.0, 25.1, 25.9, 27.0, 29.2, 29.7, 30.0, 30.2, 33.9,
35.2, 37.3, 41.0, 42.6, 43.4, 46.0, 46.2, 47.6, 47.7, 48.9, 50.2, 54.4, 55.4,
59.6, 84.6, 85.9, 109.5, 126.8, 127.7, 128.8, 144.5,150.6, 209.4; ESI-MS
m/z: 813.4 [MþH]þ; HRMS calcd for C55H77O3Si [MþH]þ: 813.5642,
found: 813.5636.

J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
7979
concentrated in vacuo. The crude product was purified by column
chromatography (petroleum ether/ethyl acetate, 10:1) to give
sodium sulfate, filtered, and evaporated under reduced pressure.
The crude product was purified by column chromatography
2
a
,3
b
-diacetyloxy-lup-20(29)-en-28-ol (18) as
a
white solid
(petroleum ether/ethyl acetate, 4:1) to give 2a,3b-dihydroxylup-
(335 mg, 76% from 17). Mp 142–144 ꢀC; IR (film, cmꢂ1) 3551, 3352,
20(29)-en-28-oic acid (3) as a white solid (82 mg, 97%). Mp 310–314 ꢀC
20
20
2944, 2868, 1741, 1641, 1454, 1393, 1369, 1252, 1233, 1037, 882, 737,
(lit.23 mp 275–278 ꢀC); [
a
]
ꢂ2.7 (c 0.39, pyridine) (lit.23
[a]
D
D
704; 1H NMR (CDCl3)
d
0.88 (s, 3H), 0.89 (s, 3H), 0.97 (s, 6H), 1.02 (s,
ꢂ4.0 (c 1.0, pyridine)); IR (KBr, cmꢂ1) 3472, 2944, 2871, 2376, 1701,
3H), 1.67 (s, 3H), 1.96 (s, 3H), 2.05 (s, 3H), 2.34–2.43 (m, 1H), 3.33
1685, 1560, 1458, 1375, 1221, 1182, 1076, 1047, 964, 885; 1H NMR
and 3.78 (d, J¼10.7 Hz, each 1H), 4.57 and 4.68 (br s, each 1H), 4.72
(C5D5N) d 0.92 (s, 3H), 1.06 (s, 9H), 1.26 (s, 3H), 1.79 (s, 3H), 1.87–
1.95 (m, 2H), 2.23–2.34 (m, 3H), 2.61–2.78 (m, 2H), 3.39 (d,
(d, J¼10.4 Hz, 1H), 5.04–5.13 (m, 1H); 13C NMR (CDCl3)
d
14.7, 16.0,
17.2, 17.4, 18.2, 19.1, 20.9, 21.0, 21.1, 25.1, 27.0, 28.3, 29.2, 29.8, 34.0,
34.1, 37.3, 38.4, 39.4, 41.0, 42.8, 44.3, 47.8, 48.8, 50.4, 55.1, 60.6, 70.3,
80.7, 109.8, 127.7, 150.3, 170.5, 170.8; ESI-MS m/z: 560.6 [MþNH4]þ;
HRMS calcd for C34H54NaO5 [MþNa]þ: 565.3868, found: 565.3863.
J¼9.4 Hz, 1H), 3.48–3.57 (m, 1H), 4.05–4.13 (m, 1H), 4.77 and 4.93
(d, J¼2.2 Hz, each 1H); 13C NMR (C5D5N)
d 14.9, 16.5, 17.4, 17.7, 18.8,
19.5, 21.4, 26.1, 29.2, 30.2, 31.2, 32.9, 34.8, 37.6, 38.6, 38.8, 39.9,
41.2, 42.9, 47.8, 48.2, 49.8, 51.0, 56.1, 56.7, 68.9, 83.8, 109.9, 151.3,
179.1; ESI-MS m/z: 471.4 [MꢂH]ꢂ; HRMS calcd for C30H48NaO4
[MþNa]þ: 495.3450, found: 495.3445.
4.8. 2a,3b-Diacetyloxy-lup-20(29)-en-28-aldehyde (19)
To a solution of compound 18 (180 mg, 0.33 mmol) in CH2Cl2
was added PCC (142 mg, 0.66 mmol), the reaction mixture was
stirred at rt for 40 min. Silica gel was poured into the reaction
mixture and the solvent was evaporated under reduced pressure to
dryness. The residue was purified by column chromatography on
4.11. 2a,3b,28-Lup-20(29)-en-triol (4)
To a solution of 17 (80 mg, 0.11 mmol) in absolute ethanol
(5 mL) was added PPTS (114 mg, 0.46 mmol). The reaction mixture
was stirred at 70 ꢀC for 8 h. After cooling to room temperature, the
solvent was evaporated in vacuo. Ice water (20 mL) was added to
the residue, and extracted with ethyl acetate (3ꢁ25 mL). The
combined extract was washed with saturated sodium bicarbonate
solution and brine, dried over anhydrous sodium sulfate, filtered,
and concentrated in vacuo. The crude product was purified by
column chromatography (petroleum ether/ethyl acetate, 3:1) to
silica gel (petroleum ether/ethyl acetate, 19:1) to give 2a,3b-diac-
etyloxy-lup-20(29)-en-28-aldehyde (19) as a white solid (173 mg,
96%). Mp 105–109 ꢀC; IR (film, cmꢂ1) 3435, 2945, 2868, 1741, 1641,
1453, 1369, 1250, 1041, 737; 1H NMR (CDCl3)
d 0.87 (s, 3H), 0.88 (s,
3H), 0.91 (s, 3H), 0.97 (s, 6H), 1.69 (s, 3H), 1.96 (s, 3H), 2.04 (s, 3H),
2.85–2.87 (m, 1H), 4.62 and 4.75 (br s, each 1H), 4.72 (d, J¼10.4 Hz,
1H), 5.04–5.13 (m, 1H), 9.67 (s, 1H); 13C NMR (CDCl3)
d
14.2, 15.9,
give 2a,3b,28-lup-20(29)-en-triol (4) as a white solid (52 mg, 99%).
20
17.2, 17.4, 18.2, 19.0, 20.8, 20.9, 21.0, 25.4, 28.3, 28.8, 29.2, 29.9, 33.2,
34.2, 38.4, 39.4, 40.9, 42.6, 44.3, 47.5, 48.1, 50.4, 55.1, 59.3, 70.2, 80.7,
110.2, 149.6, 170.7, 206.4; ESI-MS m/z: 563.6 [MþNa]þ; HRMS calcd
for C34H52NaO5 [MþNa]þ: 563.3712, found: 563.3707.
Mp 254–256 ꢀC (lit.12 mp 258–260 ꢀC); [
a
]
D
þ10.5 (c 0.24, CHCl3)
20
(lit.12
[
a]
þ11 (c 0.59, CHCl3–MeOH 9:1)); IR (film, cmꢂ1) 2918,
D
2861, 2398, 2298, 1726, 1641, 1459, 1363, 1260, 1100, 1026, 880,
745, 571; 1H NMR (CDCl3)
d 0.83 (s, 3H), 0.93 (s, 3H), 1.01 (s, 3H),
1.04 (s, 3H), 1.06 (s, 3H), 1.71 (s, 3H), 1.92–2.07 (m, 4H), 2.40–2.42
(m, 1H), 3.00 (d, J¼9.5 Hz, 1H), 3.37 (d, J¼10.8 Hz, 1H), 3.82 (dd,
J¼1.7, 10.8 Hz, 1H), 3.65–3.70 (m, 1H), 4.61 (dd, J¼1.4, 2.2 Hz, 1H),
4.9. 2a,3b-Diacetyloxy-lup-20(29)-en-28-oic acid (20)
Compound 19 (130 mg, 0.24 mmol) was dissolved in 7.5 mL of
tert-butyl alcohol, 1.5 mL of distilled THF, and 2.25 mL of 2-methyl-
2-butene. The solution was stirred and cooled in an ice-bath. Then
5.4 mL of freshly prepared aqueous solution of NaH2PO4/NaClO2
(378.7 mg/291 mg) was slowly added to the solution and the
mixture was stirred for 15 min. The mixture was then raised to rt
and stirred for 1 h. Finally, the mixture was poured into 10 mL of
saturated solution of NH4Cl and extracted with CH2Cl2. The com-
bined extract was washed with saturated sodium bicarbonate so-
lution and brine, dried over anhydrous sodium sulfate, filtered, and
concentrated in vacuo. The crude product was purified by column
chromatography (petroleum ether/ethyl acetate, 93:7) to give
4.71 (d, J¼2.2 Hz, 1H); 13C NMR (CDCl3)
d 14.8, 16.1, 16.5, 17.4, 18.4,
19.1, 21.0, 25.3, 27.2, 28.5, 29.3, 29.9, 34.0, 34.3, 37.4, 39.2, 41.1,
42.9, 46.9, 47.9, 48.9, 50.5, 53.6, 60.7, 69.3, 77.2, 84.0, 109.7, 150.4;
ESI-MS m/z: 939.8 [2MþNa]þ; HRMS calcd for C30H50O3: 458.3760,
found: 458.3764.
4.12. 28-Trityl-2a, 3a-dihydroxylup-20(29)-en-28-ol (21)
To a solution of 11 (1.76 g, 2.53 mmol) in dry isopropanol were
added 20 mL of fresh prepared aluminum isopropoxide
(1.54 mmol/ml) in dry isopropanol and cat. amount of aluminum
chloride, the reaction mixture was stirred at 110 ꢀC for 16 h. The
reaction mixture was cooled to rt, the solvent was removed in
vacuo, and the pH was adjusted to 3.0 by 1 N HCl at 0 ꢀC. The
mixture was extracted with ethyl acetate (4ꢁ40 mL). The com-
bined extract was washed with saturated sodium bicarbonate
solution and brine, dried over anhydrous sodium sulfate, filtered,
and the solvent was removed under reduced pressure. The crude
product was purified by column chromatography (petroleum
ether/ethyl acetate, 5:1) to give diol 21 as a white solid (1.06 g,
60%). Mp 179–181 ꢀC; IR (film, cmꢂ1) 3430, 2941, 2868, 2360, 2337,
2
a b-diacetyloxy-lup-20(29)-en-28-oic acid (20) as a white solid
,3
(110 mg, 82%). Mp 240–243 ꢀC; IR (film, cmꢂ1) 2947, 2869, 1741,
1694, 1454, 1369, 1250, 1041, 882, 738; 1H NMR (CDCl3)
0.88 (s,
d
6H), 0.94 (s, 3H), 0.97 (s, 6H), 1.74 (s, 3H), 1.97 (s, 3H), 2.05 (s, 3H),
2.12–2.30 (m, 3H), 2.97–3.04 (m, 1H), 4.61 and 4.74 (br s, each 1H),
4.73 (d, J¼10.2 Hz, 1H), 5.05–5.14 (m, 1H); 13C NMR (CDCl3)
d 14.6,
16.0, 17.2, 17.4, 18.2, 19.4, 20.9, 21.1, 25.4, 28.3, 29.7, 30.6, 32.2, 34.2,
37.0, 38.4, 39.4, 40.8, 42.5, 44.3, 46.9, 49.3, 50.5, 55.1, 56.3, 70.3,
80.7, 109.8, 150.2, 170.5, 170.8, 180.6; ESI-MS m/z: 555.3 [MꢂH]ꢂ;
HRMS calcd for C34H52NaO6 [MþNa]þ: 579.3661, found: 579.3656.
1555, 1453, 1063, 1036, 759, 704; 1H NMR (CDCl3)
d 0.52 (s, 3H),
0.82 (s, 3H), 0.84 (s, 3H), 0.92 (s, 3H), 1.00 (s, 3H), 1.64 (s, 3H), 2.18–
2.22 (m, 4H), 2.92 and 3.14 (d, J¼8.8 Hz, each 1H), 3.41 (d,
J¼2.5 Hz, 1H), 3.95 (m, 1H), 4.53 and 4.59 (br s, each 1H), 7.21–7.34
4.10. 2a,3b-Dihydroxylup-20(29)-en-28-oic acid (3)
Compound 20 (100 mg, 0.18 mmol) was dissolved in 5 mL of
MeOH, then 1 mL of 4 N NaOH aq was added dropwise. The re-
action mixture was stirred at rt for 2 h. The solvent was removed
in vacuo, and the pH was adjusted to 3.0 by 1 N HCl at 0 ꢀC. The
mixture was extracted with ethyl acetate (4ꢁ20 mL). The com-
bined extract was washed with brine, dried over anhydrous
(m, 9H), 7.49–7.51 (m, 6H); 13C NMR (CDCl3)
d 14.8, 15.9, 17.0, 18.0,
19.1, 20.7, 21.6, 25.1, 26.9, 28.5, 29.9, 30.2, 33.9, 35.2, 37.2, 38.3,
38.5, 40.8, 42.1, 42.6, 47.6, 47.8, 48.1, 48.9, 50.0, 59.6, 66.7, 79.0,
85.9, 109.4, 126.8, 127.7, 128.8, 144.5, 150.8; ESI-MS m/z: 699.5
[MꢂH]ꢂ; HRMS calcd for C49H64NaO3 [MþNa]þ: 723.4753, found:
723.4748.

7980
J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
4.13. 2
a
,3
a
,28-Lup-20(29)-en-triol (6)
71.3, 109.5, 126.8, 127.7, 128.8, 144.5, 150.7, 214.0; ESI-MS m/z: 813.4
[MþH]þ; HRMS calcd for C55H77O3Si [MþH]þ: 813.5642, found:
To a solution of 21 (70 mg, 0.1 mmol) in absolute ethanol (2 mL)
was added PPTS (99.8 mg, 0.4 mmol). The reaction mixture was
stirred at 70 ꢀC for 8 h. After cooling to room temperature, the
solvent was evaporated in vacuo. Ice water (20 mL) was added to
the residue, and extracted with ethyl acetate (3ꢁ20 mL). The
combined extract was washed with saturated sodium bicarbonate
solution and brine, dried over anhydrous sodium sulfate, filtered,
and concentrated in vacuo. The crude product was purified by
column chromatography (petroleum ether/ethyl acetate, 5:1) to
813.5636.
4.16. 28-Trityl-2
a-(tert-butyldimethylsilyloxy)-3b-hydroxy-
lup-20(29)-en-28-ol (24)
Compound 23 (222 mg, 0.27 mmol) was dissolved in THF
(5 mL) and EtOH (1 mL), the solution was cooled to 0 ꢀC, then
NaBH4 (14.4 mg, 0.38 mmol) was added, and the reaction mixture
was stirred for 5 h. The reaction was quenched by 1 N HCl at 0 ꢀC,
then organic solvent was removed under reduced pressure. The
mixture was extracted with ethyl acetate. The organic layers were
combined and washed with brine, dried over anhydrous sodium
sulfate, filtered, and the solvent was removed under reduced
pressure, the residue was purified by column chromatography
give 2
a,3a,28-lup-20(29)-en-triol (6) as a white solid (38.8 mg,
20
85%). Mp 242–244 ꢀC; [
a
]
þ14.4 (c 0.20, CHCl3); IR (film, cmꢂ1
)
D
3392, 2946, 2870, 1455.9, 1375, 1032, 993.3, 884; 1H NMR (CDCl3)
d
0.84 (s, 3H), 0.93 (s, 3H), 0.99 (s, 3H), 1.00 (s, 3H), 1.01 (s, 3H), 1.68
(s, 3H), 2.34–2.43 (m, 1H), 3.34 (d, J¼10.9 Hz, 1H), 3.79 (dd, J¼10.9,
1.4 Hz,1H), 3.41 (d, J¼2.7 Hz,1H), 3.94–4.00 (m,1H), 4.58 (dd, J¼2.0,
(petroleum ether/ethyl acetate, 150:1) to give 28-trityl-2a-(tert-
1.3 Hz, 1H), 4.68 (d, J¼2.0 Hz, 1H); 13C NMR (CDCl3)
d 14.8, 16.0, 17.1,
butyldimethylsilyloxy)-3b-hydroxy-lup-20(29)-en-28-ol (24) as
a white solid (209 mg, 94%). Mp 250–251 ꢀC; IR (film, cmꢂ1) 3594,
2947, 2859, 1489, 1448, 1252, 1107, 1067, 882, 837, 775, 705, 631;
18.0, 19.1, 20.8, 21.6, 25.1, 27.0, 28.4, 29.2, 29.8, 33.96, 34.00, 37.2,
38.3, 38.6, 41.1, 42.2, 42.8, 47.8, 48.2, 48.7, 50.1, 60.6, 66.6, 79.0,
109.7, 150.4; ESI-MS m/z: 457.4 [MꢂH]ꢂ; HRMS calcd for C30H50O3:
458.3760, found: 458.3761.
1H NMR (CDCl3)
d 0.05 (s, 3H), 0.07 (s, 3H), 0.51 (s, 3H), 0.80 (s,
3H), 0.81 (s, 3H), 0.87 (s, 3H), 0.88 (s, 9H), 1.01 (s, 3H), 1.64 (s, 3H),
2.17–2.24 (m, 3H), 2.91 and 3.14 (d, J¼9.0 Hz, each 1H), 2.95 (d,
J¼9.3 Hz, 1H), 3.61–3.69 (m, 1H), 4.53 and 4.58 (br s, each 1H),
4.14. 28-Trityl-2
lup-20(29)-en-28-ol (22)
a-(tert-butyldimethylsilyloxy)-3a-hydroxy-
7.20–7.50 (m, 15H); 13C NMR (CDCl3)
d 14.7, 15.6, 16.0, 16.6, 17.3,
18.0, 18.1, 19.1, 20.9, 25.2, 25.9, 26.9, 28.6, 30.0, 30.2, 34.1, 35.2,
37.2, 38.6, 38.8, 40.7, 42.6, 47.4, 47.6, 47.8, 48.9, 50.1, 55.3, 59.6,
71.0, 83.2, 109.3, 126.8, 127.7, 128.8, 144.5, 150.9; ESI-MS m/z: 813.4
[MꢂH]ꢂ; HRMS calcd for C51H69O3Si [Mꢂt-Bu]: 757.5016, found:
757.5021.
Compound 21 (300 mg, 0.43 mmol) was dissolved in 8 mL DMF,
then imidazole (234.2 mg, 3.44 mmol) and TBSCl (259.1 mg,
1.72 mmol) were added, the reaction mixture was stirred at rt for
3 h, the reaction was quenched by pouring 30 mL ice water to the
reaction mixture, then EtOAc extracted four times, the combined
organic layer was washed by brine and dried over anhydrous so-
dium sulfate, the residue was purified by column chromatography
4.17. 28-Trityl-2
a,3b-dihydroxy-lup-20(29)-en-28-ol (17)
(prepared from 24)
(petroleum ether/ethyl acetate, 20:1) to give 28-trityl-2a-(tert-
butyldimethylsilyloxy)-3a-hydroxy-lup-20(29)-en-28-ol (22) as
a white solid (320 mg, 92%). Mp 264–265 ꢀC; IR (film, cmꢂ1) 3587,
2948, 2860, 1641, 1591, 1489, 1448, 1375, 1255, 1065, 997, 901, 836,
Compound 24 (110 mg, 0.135 mmol) was dissolved in THF
(5 mL) and one drop of water was added, then 0.4 mL of TBAF so-
lution (1 M) was added to the solution, the reaction mixture was
heated to refluxing, and maintained for 2 h, then the solvent was
removed directly, and the residue was purified directly by column
chromatography (petroleum ether/ethyl acetate, 4:1) to give 28-
775, 705, 631, 583; 1H NMR (CDCl3)
d 0.04 (s, 3H), 0.05 (s, 3H), 0.50
(s, 3H), 0.78 (s, 3H), 0.81 (s, 3H), 0.87 (s, 9H), 0.89 (s, 3H), 1.00 (s,
3H), 1.63 (s, 3H), 2.13–2.22 (m, 3H), 2.45 (br s, 1H), 2.90 and 3.14 (d,
J¼8.9 Hz, each 1H), 3.25 (d, J¼2.7 Hz, 1H), 3.93–4.00 (m, 1H), 4.51
(br s, 1H), 4.57 (d, J¼2.3 Hz, 1H), 7.22–7.50 (m, 15H); 13C NMR
trityl-2
a,3b-dihydroxy-lup-20(29)-en-28-ol (17) as a white solid
(94 mg, 99%).
(CDCl3)
d 14.8, 15.9, 17.2, 17.9, 18.0, 19.0, 20.7, 21.8, 25.1, 25.8, 26.9,
28.4, 29.9, 30.2, 33.9, 35.2, 37.2, 37.8, 38.3, 40.8, 42.2, 42.6, 47.6, 47.8,
48.9, 49.9, 59.6, 68.1, 79.1, 85.9, 109.3, 126.8, 127.7, 128.8, 144.5,
150.9; ESI-MS m/z: 813.5 [MꢂH]ꢂ; HRMS calcd for C55H79O3Si
[MþH]þ: 815.5798, found: 815.5793.
4.18. 2b,3b,28-Lup-20(29)-en-triol (10)
To a solution of 2517 (320 mg, 0.46 mmol) in absolute ethanol
(5 mL) was added PPTS (456 mg, 1.82 mmol). The reaction mixture
was stirred at 70 ꢀC for 8 h. After cooling to room temperature, the
solvent was evaporated in vacuo. Ice water (50 mL) was added to
the residue, and extracted with ethyl acetate (3ꢁ35 mL). The
combined extract was washed with brine, dried over anhydrous
sodium sulfate, filtered, and concentrated in vacuo. The crude
product was purified by column chromatography (petroleum
4.15. 28-Trityl-2a-(tert-butyldimethylsilyloxy)-lup-20(29)-
en-28-ol-3-one (23)
Compound 22 (274 mg, 0.34 mmol) was dissolved in dichloro-
methane (5 mL) and cooled to 0 ꢀC, then PCC (88 mg, 0.41 mmol)
was added. Then reaction mixture was stirred overnight. The re-
action mixture was filtered, and the solvent was removed under
reduced pressure, the residue was purified by column chromatog-
ether/ethyl acetate, 3:1) to give 2
as a white solid (205 mg, 98%). Mp 194–195 ꢀC; [
0.29, CHCl3); IR (film, cmꢂ1) 3353, 2938, 1742, 1609, 1509, 1447,
1364, 1219, 1031, 877, 757; 1H NMR (CDCl3)
0.97 (s, 3H), 0.98 (s,
b,3b,28-lup-20(29)-en-triol (10)
20
a
]
þ32.6 (c
D
raphy (petroleum ether/ethyl acetate, 150:1) to give 28-trityl-2a-
(tert-butyldimethylsilyloxy)-lup-20(29)-en-28-ol-3-one (23) as
a white solid (235 mg, 86%). Mp 145–146 ꢀC; IR (film, cmꢂ1) 2948,
2861, 1722, 1489, 1449, 1374, 1250, 1121, 1064, 882, 837, 775, 705,
d
3H), 0.99 (s, 3H), 1.04 (s, 3H), 1.14 (s, 3H), 1.68 (s, 3H), 1.71–1.96 (m,
3H), 2.04–2.17 (m, 3H), 2.34–2.40 (m, 1H), 3.19 (br s, 1H), 3.33 and
3.80 (d, J¼10.8 Hz, each 1H), 4.08 (br s, 1H), 4.58 and 4.69 (br s,
632; 1H NMR (CDCl3)
(s, 3H), 0.88 (s, 9H), 1.03 (s, 3H), 1.06 (s, 3H), 1.08 (s, 3H), 1.63 (s, 3H),
d
ꢂ0.01 (s, 6H), 0.11 (s, 3H), 0.56 (s, 3H), 0.86
each 1H); 13C NMR (CDCl3)
d 14.7, 16.0, 17.1, 18.1, 19.1, 21.0, 25.3,
2.16–2.24 (m, 4H), 2.92 and 3.13 (d, J¼9.0 Hz, each 1H), 4.49–4.58
27.0, 29.2, 29.6, 29.8, 34.0, 34.2, 36.9, 37.3, 38.2, 41.1, 42.9, 44.5,
47.8, 48.8, 50.9, 55.3, 60.6, 71.2, 78.5, 109.7, 150.4; ESI-MS m/z:
481.4 [MþNa]þ; HRMS calcd for C30H50O3: 458.3760, found:
458.3762.
(m, 3H), 7.20–7.50 (m,15H); 13C NMR (CDCl3)
d
ꢂ5.5, ꢂ4.6,14.6,16.1,
16.8, 18.5, 19.0, 19.2, 21.1, 21.6, 25.1, 25.2, 25.8, 26.9, 29.9, 30.2, 34.0,
35.2, 37.3, 37.9, 40.8, 42.6, 47.6, 47.8, 48.3, 48.9, 50.0, 50.4, 56.7, 59.6,

J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
7981
4.19. 28-Trityl-2
lup-20(29)-en-28-ol (14)
b
-(tert-butyldimethylsilyloxy)-3
b
-hydroxy-
(s, 3H), 0.87 (s, 12H), 0.89 (s, 3H), 0.96 (s, 3H), 1.00 (s, 3H), 1.65 (s,
3H), 2.16–2.24 (m, 4H), 2.91 and 3.13 (d, J¼8.8 Hz, each 1H), 3.54 (d,
J¼8.9 Hz, 1H), 3.68–3.73 (m, 1H), 4.54 and 4.59 (d, J¼2.1 Hz, each
Compound 25 (1.57 g, 2.2 mmol) was dissolved in 28 mL of DMF,
then imidazole (1.20 g, 17.6 mmol) and tert-butyldimethysilyl
chloride (1.36 g, 9.02 mmol) were added. The reaction mixture was
stirred at 40 ꢀC for 2 h. The mixture was cooled to rt, and then
10 mL of ice water was added to the reaction mixture, the mixture
was extracted with ethyl acetate (4ꢁ30 mL). The combined extract
was washed with saturated sodium bicarbonate solution and brine,
dried over anhydrous sodium sulfate, filtered, and the solvent was
removed under reduced pressure. The crude product was purified
by column chromatography (petroleum ether/ethyl acetate, 160:1)
1H), 7.22–7.50 (m, 15H); 13C NMR (CDCl3)
d 14.7, 15.7, 18.0,19.2,19.4,
20.7, 21.4, 23.0, 24.8, 25.4, 25.8, 26.9, 30.0, 30.1, 33.4, 35.2, 37.1, 37.5,
37.7, 40.8, 42.6, 47.4, 47.6, 47.8, 48.9, 50.4, 51.1, 59.6, 71.3, 85.9, 109.3,
126.8, 127.7, 128.8, 144.5, 150.9; ESI-MS m/z: 813.8 [MꢂH]ꢂ; HRMS
calcd for C55H78NaO3Si [MþNa]þ: 837.5617, found: 837.5612.
4.22. 28-Trityl-2b,3a-dihydroxy-lup-20(29)-en-28-ol (28)
Compound 27 (1.42 g, 1.74 mmol) was dissolved in THF (95 mL)
and 14 drops of water were added, then TBAF (0.68 g, 2.60 mmol) in
THF was added to the solution, the reaction mixture was heated to
reflux, and maintained for 2 h, then the solvent was removed
directly, and the residue was purified directly by column chroma-
tography (petroleum ether/ethyl acetate, 4:1) to give 28-trityl-
to give 28-trityl-2b-(tert-butyldimethylsilyloxy)-3b-hydroxy-lup-
20(29)-en-28-ol (14) as a white solid (1.49 g, 82%). Mp 128–130 ꢀC;
IR (film, cmꢂ1) 3580, 2949, 2854,1644,1489, 1448,1375, 1254,1084,
1065, 1035, 976, 897, 836, 775, 765, 706, 631; 1H NMR (CDCl3)
d 0.05
(s, 3H), 0.07 (s, 3H), 0.49 (s, 3H), 0.88 (s, 9H), 0.89 (s, 3H), 0.92 (s,
3H), 0.96 (s, 3H), 1.03 (s, 3H), 1.62 (s, 3H), 1.96–2.21 (m, 5H), 2.91
and 3.10 (d, J¼8.7 Hz, each 1H), 3.01 (br s, 1H), 4.02–4.03 (m, 1H),
4.51 (br s, 1H), 4.57 (d, J¼1.8 Hz, 1H), 7.22–7.32 (m, 9H), 7.47–7.50
2
b
,3
a
-dihydroxy-lup-20(29)-en-28-ol (28) as
a white solid
(1.10 mg, 90%). Mp 165–167 ꢀC; IR (film, cmꢂ1) 3400, 2939, 2868,
1637, 1489, 1448, 1374, 1264, 1063, 987, 883, 764, 742, 706, 631; 1H
NMR (CDCl3)
0.98 (s, 3H), 1.63 (s, 3H), 2.16–2.20 (m, 3H), 2.90 and 3.12 (d,
d 0.50 (s, 3H), 0.88 (s, 3H), 0.89 (s, 3H), 0.93 (s, 3H),
(m, 6H); 13C NMR (CDCl3)
d
ꢂ5.2, ꢂ3.9, 14.9, 16.2, 17.0, 17.2, 18.3,
19.3, 21.1, 25.4, 26.1, 27.0, 29.8, 30.0, 30.1, 30.4, 34.3, 35.5, 37.1, 37.5,
38.7, 40.9, 42.8, 45.3, 47.8, 48.0, 49.1, 50.8, 55.5, 59.8, 72.4, 78.4,
109.6, 127.1, 128.0, 129.0, 144.8, 151.1; ESI-MS m/z: 813.9 [MꢂH]ꢂ;
HRMS calcd for C55H78NaO3Si [MþNa]þ: 837.5617, found: 837.5612.
J¼8.8 Hz, each 1H), 3.59–3.70 (m, 2H), 4.52 and 4.58 (br s, each 1H),
7.20–7.50 (m, 15H); 13C NMR (CDCl3)
d 14.7, 15.6, 19.1, 19.9, 21.6,
23.2, 23.4, 25.4, 26.9, 30.0, 30.1, 33.3, 35.2, 37.59, 37.63, 40.8, 42.6,
47.6, 47.8, 48.0, 48.9, 51.0, 51.1, 59.6, 69.1, 78.1, 109.4, 126.8, 127.7,
128.8, 129.7, 144.5, 150.7; ESI-MS m/z: 699.5 [MꢂH]ꢂ; HRMS calcd
for C49H64NaO3 [MþNa]þ: 723.4753, found: 723.4748.
4.20. 28-Trityl-2
28-ol-3-one (26)
b-(tert-butyldimethylsilyloxy)-lup-20(29)-en-
4.23. 2b,3a,28-Lup-20(29)-en-triol (8)
To a solution of 14 (1.8 g, 2.2 mmol) in dichloromethane was
added PCC (0.57 g, 2.65 mol), the reaction mixture was stirred at
0 ꢀC for 12 h, the mixture was filtered and concentrated in vacuo,
the residue was purified directly by column chromatography (pe-
To a solution of 28 (160 mg, 0.23 mmol) in absolute ethanol
(5 mL) was added PPTS (228 mg, 0.91 mmol). The reaction mixture
was stirred at 70 ꢀC for 8 h. After cooling to room temperature, the
solvent was evaporated in vacuo. Ice water (20 mL) was added to
the residue, and extracted with ethyl acetate (3ꢁ25 mL). The
combined extract was washed with brine, dried over anhydrous
sodium sulfate, filtered, and concentrated in vacuo. The crude
product was purified by column chromatography (petroleum ether/
troleum ether/ethyl acetate, 150:1) to give 28-trityl-2b-(tert-
butyldimethylsilyloxy)-lup-20(29)-en-28-ol-3-one (26) as a white
solid (1.40 g, 78%). Mp 162–164 ꢀC; IR (film, cmꢂ1) 2949, 2863,1726,
1448, 1385, 1250, 1064, 991, 837, 741, 706, 632; 1H NMR (CDCl3)
d
ꢂ0.01 (s, 3H), 0.12 (s, 3H), 0.49 (s, 3H), 0.68 (s, 3H), 0.87 (s, 9H),
0.95 (s, 3H), 1.05 (s, 3H), 1.07 (s, 3H), 1.64 (s, 3H), 2.00–2.07 (m, 1H),
2.14–2.25 (m, 3H), 2.91 and 3.12 (d, J¼8.8 Hz, each 1H), 4.52 and
4.58 (br s, each 1H), 4.62–4.69 (m, 1H), 7.19–7.49 (m, 15H); 13C NMR
ethyl acetate, 3:1) to give 2b,3a,28-lup-20(29)-en-triol (8) as
20
a white solid (101.6 mg, 97%). Mp 230–232 ꢀC; [
a
]
þ46.6 (c 0.259,
D
CHCl3); IR (KBr, cmꢂ1) 3373, 2934, 2865, 2391, 2292, 1741, 1723,
1640, 1453, 1410, 1387, 1372, 1240, 1108, 1077, 1043, 1023, 978, 880,
(CDCl3)
d 14.7, 15.2, 18.6, 19.1, 19.4, 20.0, 21.9, 25.4, 25.8, 26.9, 29.7,
29.9, 30.1, 32.8, 35.2, 37.1, 37.6, 40.5, 42.6, 46.2, 47.6, 47.7, 48.9, 49.8,
51.8, 52.0, 59.6, 70.9, 109.4, 126.8, 127.7, 128.8, 144.4, 150.7; ESI-MS
m/z: 813.4 [MþH]þ; HRMS calcd for C55H77O3Si [MþH]þ: 813.5642,
found: 813.5636.
560, 544, 517; 1H NMR (C5D5N)
d 0.98 (s, 3H), 1.00 (s, 3H), 1.18 (s,
3H), 1.23 (s, 3H), 1.26 (s, 3H), 1.75 (s, 3H), 2.00–2.17 (m, 2H), 2.36–
2.44 (m, 2H), 2.55–2.64 (m, 1H), 3.64 and 4.07 (d, J¼10.8 Hz, each
1H), 3.98 (d, J¼8.4 Hz,1H), 4.24–4.31 (m,1H), 4.72 (br s,1H), 4.87 (d,
J¼2.3 Hz, 1H); 13C NMR (C5D5N)
d 14.9, 16.0, 19.3, 19.9, 20.9, 21.8,
4.21. 28-Trityl-2
b
-(tert-butyldimethylsilyloxy)-3
a
-hydroxy-
23.5, 25.9, 26.2, 27.5, 30.0, 30.4, 34.1, 34.9, 37.8, 37.9, 38.1, 41.4, 43.1,
47.2, 48.3, 48.6, 49.1, 50.9, 51.4, 59.4, 70.3, 78.2, 109.9, 151.2; ESI-MS
m/z: 476.4 [MþNH4]þ; HRMS calcd for C30H50NaO3 [MþNa]þ:
481.3657, found: 481.3652.
lup-20(29)-en-28-ol (27)
To a solution of 26 (600 mg, 0.74 mmol) in dry isopropanol were
added 8 mL of fresh prepared aluminum isopropoxide (1.54 mmol/
ml) in dry isopropanol and cat. amount of aluminum chloride, the
reaction mixture was stirred at 110 ꢀC for 18 h. The reaction mixture
was cooled to rt, the solvent was removed in vacuo, and the pH was
adjusted to 3.0 by 1 N HCl at 0 ꢀC. The mixture was extracted with
ethyl acetate (4ꢁ40 mL). The combined extract was washed with
saturated sodium bicarbonate solution and brine, dried over an-
hydrous sodium sulfate, filtered, and the solvent was removed
under reduced pressure. The crude product was purified by column
chromatography (petroleum ether/ethyl acetate, 150:1) to give 28-
4.24. 2a,3a-Diacetyloxy-lup-20(29)-en-28-ol (29)
To a solution of 21 (500 mg, 0.713 mmol) in dry pyridine (3 mL)
was added Ac2O (1 mL), the reaction mixture was stirred at rt
overnight. After cooling to 0 ꢀC, 1 N HCl (25 mL) was added into the
reaction mixture. The mixture was extracted with ethyl acetate
(3ꢁ30 mL). The combined extract was washed with saturated so-
dium bicarbonate solution and brine, dried over anhydrous sodium
sulfate, filtered, and concentrated in vacuo. The crude product was
used directly without further purification. The crude residue was
dissolved in absolute ethanol (10 mL) and was then added PPTS
(637 mg, 2.55 mmol). The reaction mixture was stirred at 70 ꢀC for
8 h. After cooling to room temperature, the solvent was evaporated
trityl-2b-(tert-butyldimethylsilyloxy)-3a-hydroxy-lup-20(29)-en-28-
ol (27) as a white solid (318.8 mg, 53%). Mp 136–138 ꢀC; IR (film,
cmꢂ1) 3395, 2946, 2861, 1722, 1594, 1449, 1384, 1250, 1087, 1064,
835, 774, 705, 632; 1H NMR (CDCl3)
d 0.05 (s, 3H), 0.06 (s, 3H), 0.50

7982
J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
in vacuo. Ice water (15 mL) was added to the residue, and extracted
with ethyl acetate (3ꢁ20 mL). The combined extract was washed
with saturated sodium bicarbonate solution and brine, dried over
anhydrous sodium sulfate, filtered, and concentrated in vacuo. The
crude product was purified by column chromatography (petroleum
563.6 [MþNa]þ; HRMS calcd for C34H52NaO5 [MþNa]þ: 563.3712,
found: 563.3707.
4.28. 2
b,3b-Diacetyloxy-lup-20(29)-en-28-aldehyde (33)
ether/ethyl acetate, 20:1) to give 2
28-ol (29) as a white solid (282 mg, 73%). Mp 233–235 ꢀC; IR (film,
cmꢂ1) 3551, 2943, 2870, 1745, 1453, 1374, 1252, 1035, 882, 737; 1
NMR (CDCl3) 0.86 (s, 3H), 0.95 (s, 3H), 0.97 (s, 3H),1.03 (s, 6H),1.68
a
,3
a
-diacetyloxy-lup-20(29)-en-
Following the procedure described for preparation of 32, 2b,3b-
diacetyloxy-lup-20(29)-en-28-aldehyde (33) was synthesized from
30 as a white solid (89%). Mp 227–229 ꢀC; IR (film, cmꢂ1) 2944,
2868, 1742, 1641, 1450, 1395, 1367, 1252, 1188, 1031, 883, 737; 1H
H
d
(s, 3H), 1.94 (s, 3H), 2.12 (s, 3H), 2.33–2.43 (m, 1H), 3.34 and 3.80 (d,
J¼11.1 Hz, each 1H), 4.58 (dd, J¼2.2, 1.4 Hz, 1H), 4.68 (d, J¼2.2 Hz,
1H), 4.95 (d, J¼2.3 Hz, 1H), 5.19–5.25 (m, 1H); 13C NMR (CDCl3)
NMR (CDCl3) d 0.88 (s, 3H), 0.93 (s, 3H), 0.96 (s, 3H), 1.02 (s, 3H), 1.10
(s, 3H), 1.70 (s, 3H), 2.02 (s, 3H), 2.03 (s, 3H), 2.81–2.90 (m, 1H), 4.60
(d, J¼3.9 Hz, 1H), 4.64 and 4.75 (br s, each 1H), 5.30–5.33 (m, 1H),
d
15.0, 16.0, 17.0, 17.8, 19.1, 20.8, 21.0, 21.1, 21.4, 25.1, 27.1, 27.7, 29.2,
9.67 (s, 1H); 13C NMR (CDCl3)
d 14.2, 16.1, 16.8, 17.5, 18.0, 19.0, 20.8,
29.8, 34.0, 37.2, 38.2, 38.7, 39.2, 41.2, 42.9, 47.7, 47.8, 48.8, 49.8, 50.3,
60.6, 68.4, 77.2, 109.8, 150.4, 170.4, 170.6; ESI-MS m/z: 565.4
[MþNa]þ; HRMS calcd for C34H54NaO5 [MþNa]þ: 565.3868, found:
565.3864.
20.9, 21.2, 25.5, 28.7, 29.0, 29.2, 29.9, 33.3, 34.2, 37.0, 37.4, 38.7, 41.0,
42.3, 42.8, 47.6, 48.1, 51.0, 55.2, 59.3, 69.6, 78.0, 110.2, 149.6, 170.2,
170.7, 206.5; ESI-MS m/z: 558.3 [MþNH4]þ; HRMS calcd for
C34H52O5: 540.3815, found: 540.3817.
4.29. 2
b,3a-Diacetyloxy-lup-20(29)-en-28-aldehyde (34)
4.25. 2b,3b-Diacetyloxy-lup-20(29)-en-28-ol (30)
Following the procedure described for preparation of 32, 2
b,3a-
Following the procedure described for preparation of 29, 2
b,3b-
diacetyloxy-lup-20(29)-en-28-aldehyde (34) was synthesized from
31 as a white solid (92%). Mp 188–190 ꢀC; IR (film, cmꢂ1) 2943,
2869, 1742, 1641, 1452, 1369, 1246, 1027, 884, 736; 1H NMR (CDCl3)
diacetyloxy-lup-20(29)-en-28-ol (30) was synthesized from 25 as
a white solid (72%). Mp 241–242 ꢀC; IR (KBr, cmꢂ1) 3506, 2947,
2399, 2284, 1722, 1641, 1462, 1398, 1375, 1367, 1228, 1188, 1031, 981,
895, 879, 663, 605; 1H NMR (CDCl3)
d 0.88 (s, 3H), 0.97 (s, 3H), 1.02
d
0.91 (s, 6H), 0.96 (s, 3H), 0.99 (s, 3H), 1.06 (s, 3H), 1.69 (s, 3H), 2.00
(s, 3H), 2.06 (s, 3H), 2.81–2.90 (m, 1H), 4.62 and 4.75 (br s, each 1H),
(s, 3H), 1.04 (s, 3H), 1.10 (s, 3H), 1.68 (s, 3H), 2.02 (s, 3H), 2.03 (s, 3H),
4.91–4.98 (m, 1H), 5.08 (d, J¼8.0 Hz, 1H), 9.67 (s, 1H); 13C NMR
2.34–2.42 (m, 1H), 3.33 and 3.78 (d, J¼10.9 Hz, each 1H), 4.58–4.60
(m, 2H), 4.68 (br s, 1H), 5.29–5.33 (m, 1H); 13C NMR (CDCl3)
d 14.7,
(CDCl3)
d 14.4, 15.7, 19.0, 19.6, 20.9, 21.2, 21.3, 22.3, 25.6, 26.0, 28.8,
29.2, 29.9, 33.2, 33.6, 37.0, 37.4, 38.9, 40.9, 42.8, 43.0, 47.5, 48.1, 50.7,
51.0, 59.3, 70.5, 76.0, 110.2, 149.5, 170.1, 170.3, 206.5; ESI-MS m/z:
558.3 [MþNH4]þ; HRMS calcd for C34H52NaO5 [MþNa]þ: 563.3712,
found: 563.3707.
16.1, 16.7, 17.5, 18.0, 19.0, 20.8, 21.0, 21.2, 25.1, 27.0, 29.0, 29.2, 29.7,
34.0, 34.1, 36.9, 37.2, 37.4, 41.0, 42.2, 42.9, 47.8, 48.7, 50.8, 55.2, 60.6,
69.6, 78.0, 109.8, 150.3, 170.2, 170.7; ESI-MS m/z: 565.4 [MþNa]þ;
HRMS calcd for C34H55O5 [MþH]þ: 543.4049, found: 543.4044.
4.30. 2a,3a-Diacetyloxy-lup-20(29)-en-28-oic acid (35)
4.26. 2b,3a-Diacetyloxy-lup-20(29)-en-28-ol (31)
Compound 32 (50 mg, 0.092 mmol) was dissolved in 2.9 mL
of tert-butyl alcohol, 0.58 mL of distilled THF and 0.87 mL of
2-methyl-2-butene. The solution was stirred and cooled in an ice-
bath. Then 2.8 mL of freshly prepared aqueous solution of NaH2PO4/
NaClO2 (146.2 mg/112.14 mg) was slowly added to the solution and
the mixture was stirred for 15 min. The mixture was then raised to
rt and stirred for 1 h. Finally, the mixture was poured into 5 mL of
a saturated solution of NH4Cl and extracted with CH2Cl2. The
combined extract was washed with saturated sodium bicarbonate
solution and brine, dried over anhydrous sodium sulfate, filtered,
and concentrated in vacuo. The crude product was purified by
column chromatography (petroleum ether/ethyl acetate, 93:7) to
Following the procedure described for preparation of 29, 2
b,3a-
diacetyloxy-lup-20(29)-en-28-ol (31) was synthesized from 28 as
a white solid (71%). Mp 104–106 ꢀC; IR (film, cmꢂ1) 3466, 2943,
2870, 1742, 1455, 1371, 1249, 1028, 882, 737; 1H NMR (CDCl3)
d 0.91
(s, 3H), 0.97 (s, 3H), 1.00 (s, 3H), 1.02 (s, 3H), 1.06 (s, 3H), 1.67 (s, 3H),
1.99 (s, 3H), 2.06 (s, 3H), 2.33–2.40 (m, 1H), 3.33 and 3.78 (d,
J¼10.8 Hz, each 1H), 4.57 and 4.67 (br s, each 1H), 4.91–4.98 (m,
1H), 5.08 (d, J¼8.1 Hz, 1H); 13C NMR (CDCl3)
d 14.9, 15.8, 19.0, 19.6,
21.0, 21.2, 21.4, 22.3, 25.3, 26.0, 27.0, 29.2, 29.8, 33.6, 34.0, 37.0, 37.4,
37.5, 41.0, 42.9, 43.0, 47.8, 48.8, 50.7, 50.9, 60.6, 70.6, 76.1, 109.8,
150.3, 170.1, 170.4; ESI-MS m/z: 560.6 [MþNH4]þ; HRMS calcd for
C34H54NaO5 [MþNa]þ: 565.3869, found: 565.3864.
give 2a,3a-diacetyloxy-lup-20(29)-en-28-oic acid (35) as a white
solid (42 mg, 84%). Mp 182–183 ꢀC; IR (film, cmꢂ1) 2946, 2870,
1744, 1695, 1452, 1374, 1251, 1234, 1155, 1036, 885, 737, 609; 1H
4.27. 2a,3a-Diacetyloxy-lup-20(29)-en-28-aldehyde (32)
NMR (CDCl3) d 0.86 (s, 3H), 0.94 (s, 6H), 0.96 (s, 3H),1.03 (s, 3H),1.69
To a solution of 29 (200 mg, 0.369 mmol) in CH2Cl2 was added
PCC (159 mg, 0.738 mmol), the reaction mixture was stirred at rt for
40 min. Silica gel was poured into the reaction mixture and the
solvent was evaporated under reduced pressure to dryness. The
residue was directly purified by column chromatography on silica
(s, 3H), 1.94 (s, 3H), 2.12 (s, 3H), 2.97–3.04 (m, 1H), 4.61 and 4.74 (br
s, each 1H), 4.95 (d, J¼2.4 Hz, 1H), 5.19–5.25 (m, 1H); 13C NMR
(CDCl3) d 14.9, 16.1, 17.0, 17.8, 19.4, 20.8, 20.96, 21.05, 21.4, 25.4, 27.7,
29.7, 30.6, 32.2, 34.1, 37.0, 38.2, 38.3, 38.7, 39.2, 41.0, 42.6, 46.9, 49.3,
49.8, 50.4, 56.3, 68.4, 77.4, 109.7, 150.3, 170.4, 170.6, 180.5; ESI-MS
m/z: 574.6 [MþNH4]þ. Anal. Calcd for C34H53O6: C, 73.21; H, 9.58.
Found: C, 73.21; H, 9.625.
gel (petroleum ether/ethyl acetate, 19:1) to give 2a,3a-diacetyloxy-
lup-20(29)-en-28-aldehyde (32) as a white solid (173 mg, 87%). Mp
112–114 ꢀC; IR (KBr, cmꢂ1) 3437, 2945, 2872, 2376, 1734, 1458, 1375,
1232, 1036, 1157, 1036, 953, 888, 614; 1H NMR (CDCl3)
d
0.86 (s, 3H),
4.31. 2b,3b-Diacetyloxy-lup-20(29)-en-28-oic acid (36)
0.92 (s, 3H), 0.94 (s, 3H), 0.96 (s, 3H), 1.02 (s, 6H),1.70 (s, 3H),1.94 (s,
3H), 2.12 (s, 3H), 2.82–2.91 (m, 1H), 4.63 and 4.76 (d, J¼1.7 Hz, each
1H), 4.95 (d, J¼2.4 Hz, 1H), 5.19–5.25 (m, 1H); 13C NMR (CDCl3)
Following the procedure described for preparation of 35, 2
b,3b-
diacetyloxy-lup-20(29)-en-28-oic acid (36) was synthesized from
33 as a white solid (86%). Mp 246–248 ꢀC; IR (film, cmꢂ1) 2945,
2870, 1743, 1693, 1451, 1369, 1252, 1188, 1055, 1031, 982, 885, 738;
d
14.5, 16.0, 17.0, 17.8, 19.0, 20.8, 20.96, 21.05, 21.4, 25.5, 27.7, 28.8,
29.3, 29.9, 33.2, 34.1, 38.2, 38.6, 38.7, 39.2, 41.1, 42.7, 47.5, 48.1, 49.8,
50.4, 59.3, 68.4, 77.4, 110.2, 149.6, 170.3, 170.6, 206.5; ESI-MS m/z:
1H NMR (CDCl3)
d 0.88 (s, 3H), 0.96 (s, 3H), 0.97 (s, 3H), 1.02 (s, 3H),

J. Hao et al. / Tetrahedron 65 (2009) 7975–7984
7983
20
1.10 (s, 3H), 1.69 (s, 3H), 2.02 (s, 3H), 2.03 (s, 3H), 2.17–2.30 (m, 3H),
as a white solid (96%). Mp 273–276 ꢀC; [
a]
þ31.1 (c 0.262, pyri-
D
2.97–3.03 (m, 1H), 4.60 and 4.74 (d, J¼1.9 Hz, each 1H), 4.61 (s, 1H),
dine); IR (KBr, cmꢂ1) 3523, 2941, 2391, 2291, 1709, 1458, 1375, 1176,
5.30–5.34 (m, 1H); 13C NMR (CDCl3)
d
14.6, 16.2, 16.8, 17.5, 18.0, 19.4,
1158, 1133, 1057, 1029, 875, 756; 1H NMR (C5D5N)
d 1.09 (s, 3H), 1.10
20.8, 21.0, 21.2, 25.4, 29.0, 29.6, 29.7, 30.6, 32.2, 34.2, 37.1, 37.4, 38.4,
40.9, 42.2, 42.6, 46.9, 49.3, 51.0, 55.3, 56.3, 69.7, 78.0, 109.8, 150.2,
170.2, 170.7, 180.6; ESI-MS m/z: 555.3 [MꢂH]ꢂ; HRMS calcd for
C34H52O6: 556.3764, found: 556.3763.
(s, 3H), 1.24 (s, 3H), 1.34 (s, 3H), 1.43 (s, 3H), 1.80 (s, 3H), 2.18–2.29
(m, 2H), 2.35–2.40 (m, 1H), 2.62–2.79 (m, 2H), 3.44 (d, J¼3.9 Hz,
1H), 3.53–3.58 (m, 1H), 4.43–4.44 (m, 1H), 4.77 and 4.95 (br s, each
1H); 13C NMR (C5D5N)
d 15.0, 16.5, 17.5, 18.0, 18.7, 19.5, 21.5, 26.2,
30.1, 30.2, 31.3, 33.0, 34.9, 37.5, 37.6, 38.7, 38.9, 41.3, 43.0, 45.5, 47.8,
49.9, 51.5, 56.0, 56.7, 71.5, 78.4,109.9,151.4,178.8; ESI-MS m/z: 471.4
[MꢂH]ꢂ; HRMS calcd for C34H48O4: 472.3553, found: 472.3554.
4.32. 2
b,3a-Diacetyloxy-lup-20(29)-en-28-oic acid (37)
Following the procedure described for preparation of 35, 2
b,3a-
diacetyloxy-lup-20(29)-en-28-oic acid (37) was synthesized from
34 as a white solid (88%). Mp 267–269 ꢀC; IR (film, cmꢂ1) 2944,
2870, 1742, 1694, 1641, 1455, 1370, 1247, 1028, 880, 737; 1H NMR
5. Enzyme assay
The inhibitory activity of the test compounds against rabbit
muscle glycogen phosphorylase a (GPa) was monitored using
microplate reader (BIO-RAD) based on the published method. In
brief, GPa activity was measured in the direction of glycogen syn-
thesis by the release of phosphate from glucose-1-phosphate. Each
test compound was dissolved in DMSO and diluted at different
concentrations for IC50 determination. The enzyme was added into
100 L of buffer containing 50 mM Hepes (pH¼7.2), 100 mM KCl,
2.5 mM MgCl2, 0.5 mM glucose-1-phosphate, 1 mg/mL glycogen
and the test compound in 96-well microplates (Costar). After the
addition of 150 L of 1 M HCl containing 10 mg/mL ammonium
molybdate and 0.38 mg/mL malachite green, reactions were run at
22 ꢀC for 25 min, and then the phosphate absorbance was mea-
sured at 655 nm. The IC50 values were estimated by fitting the in-
hibition data to a dose-dependent curve using a logistic derivative
equation.
(CDCl3)
d 0.91 (s, 3H), 0.93 (s, 3H), 0.96 (s, 3H), 1.00 (s, 3H), 1.06 (s,
3H), 1.68 (s, 3H), 2.00 (s, 3H), 2.06 (s, 3H), 2.16–2.30 (m, 2H), 2.96–
3.04 (m, 1H), 4.61 and 4.74 (br s, each 1H), 4.92–4.98 (m, 1H), 5.08
(d, J¼8.1 Hz, 1H); 13C NMR (CDCl3)
d 14.8, 15.8, 19.0, 19.3, 19.6, 20.9,
21.2, 21.4, 22.3, 25.5, 26.0, 29.6, 30.6, 32.2, 33.6, 36.9, 37.0, 37.4, 38.6,
40.8, 42.6, 43.0, 46.9, 49.3, 50.7, 51.0, 56.3, 70.6, 76.6, 109.8, 150.2,
170.1, 170.3; ESI-MS m/z: 555.3 [MꢂH]ꢂ; HRMS calcd for
C34H52NaO6 [MþNa]þ: 579.3662, found: 579.3656.
4.33. 2a,3a-Dihydroxylup-20(29)-en-28-oic acid (5)
Compound 35 (25 mg, 0.04 mmol) was dissolved in 5 mL of
MeOH, then 1 mL of 4 N NaOH aq was added dropwise. The reaction
mixture was stirred at rt for 2 h. The solvent was removed in vacuo,
and the pH was adjusted to 3.0 by 1 N HCl at 0 ꢀC. The mixture was
extracted with ethyl acetate (4ꢁ10 mL). The combined extract was
washed with brine, dried over anhydrous sodium sulfate, filtered,
and evaporated under reduced pressure. The crude product was
purified by column chromatography (petroleum ether/ethyl ace-
Acknowledgements
This program was financially supported by the National Natural
Science Foundation (grants 30672523 and 90713037), research
grants from the Ministry of Education (grants 706030 and
20050316008), and program for New Century Excellent Talents in
University (NCET-05-0495).
tate, 4:1) to give 2a,3a-dihydroxylup-20(29)-en-28-oic acid (5) as
a white solid (20 mg, 96%). Mp 257–259 ꢀC (lit.11 mp 298–300 ꢀC);
20
[
a]
þ18.7 (c 0.227, pyridine); IR (film, cmꢂ1) 3517, 3387, 2943,
D
2870, 1733, 1694, 1641, 1455, 1376, 1153, 1125, 1034, 881; 1H NMR
(C5D5N)
d 0.86 (s, 6H), 0.92 (s, 3H), 1.03 (s, 3H), 1.23 (s, 3H), 1.74 (s,
3H), 1.87–1.99 (m, 3H), 2.19–2.25 (m, 2H), 2.56–2.73 (m, 2H), 3.47–
Supplementary data
3.54 (m, 1H), 3.74 (d, J¼2.6 Hz, 1H), 4.26–4.30 (m, 1H), 4.74 (br s,
1H), 4.90 (d, J¼2.1 Hz, 1H); 13C NMR (C5D5N)
d 14.9, 16.5, 17.5, 18.5,
Supplementary data associated with this article can be found in
the online version, at doi:10.1016/j.tet.2009.07.047.
19.5, 21.2, 22.1, 26.1, 29.4, 30.0, 30.3, 31.2, 32.9, 34.8, 37.6, 38.6, 38.9,
41.4, 42.9, 43.3, 47.8, 48.8, 49.8, 50.8, 56.6, 66.3, 79.4, 109.9, 151.3,
178.8; ESI-MS m/z: 471.3 [MꢂH]ꢂ; HRMS calcd for C30H48NaO4:
472.3553, found: 472.3559.
References and notes
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20
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d
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b,3b-
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