Stereoselective synthesis of bioactive isosteviol derivatives as α-glucosidase inhibitors

Article abstract of DOI:10.1016/j.bmc.2009.01.017

Considerable interest has been attracted in isosteviol and its derivatives because of their large variety of pharmacological activities. In this project, a series of novel compounds containing hydroxyl, hydroxymethyl group and heteroatom-containing frameworks fused with isosteviol structure were synthesized and evaluated as α-glucosidase inhibitors, aimed at clarifying the structure-activity correlation. The results indicated that these isosteviol derivatives were capable of inhibiting in vitro α-glucosidase with moderate to good activities. Among them, indole derivative 15b exhibited the highest activities and thus may be exploitable as a lead compound for the development of potent α-glucosidase inhibitors.

Full text of DOI:10.1016/j.bmc.2009.01.017

Bioorganic & Medicinal Chemistry 17 (2009) 1464–1473
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry
journal homepage: www.elsevier.com/locate/bmc
Stereoselective synthesis of bioactive isosteviol derivatives
as a-glucosidase inhibitors
*
Ya Wu, Jing-Hua Yang, Gui-Fu Dai, Cong-Jun Liu, Guo-Qiang Tian, Wen-Yan Ma, Jing-Chao Tao
Department of Chemistry, New Drug Research & Development Center, Zhengzhou University, Zhengzhou 450052, China
a r t i c l e i n f o
a b s t r a c t
Article history:
Considerable interest has been attracted in isosteviol and its derivatives because of their large variety of
pharmacological activities. In this project, a series of novel compounds containing hydroxyl, hydroxy-
methyl group and heteroatom-containing frameworks fused with isosteviol structure were synthesized
Received 17 December 2008
Revised 8 January 2009
Accepted 9 January 2009
Available online 15 January 2009
and evaluated as
results indicated that these isosteviol derivatives were capable of inhibiting in vitro
moderate to good activities. Among them, indole derivative 15b exhibited the highest activities and thus
may be exploitable as a lead compound for the development of potent -glucosidase inhibitors.
Ó 2009 Published by Elsevier Ltd.
a
-glucosidase inhibitors, aimed at clarifying the structure–activity correlation. The
a
-glucosidase with
Keywords:
Isosteviol
1,3-Diol
a
Hydroxylation
Beckmann fragmentation
Indole
a-Glycosidase
1. Introduction
In this study, a series of novel isosteviol derivatives were syn-
thesized by a facile route, and the -glucosidase inhibition activi-
ties of the derivatives were appraised, which would be aiding in
designing and synthesizing novel stronger -glucosidase inhibitors
and clarifying the structure–activity correlation involved in the
inhibition process of -glucosidase.
a
During the last few decades, there has been widespread interest
in
only in carbohydrate digestion, but also in the processing of gluco-
proteins and glycolipids. In addition, the -glycosidase inhibitors
a-glucosidase (EC 3.2.1.20) because of its important role not
a
a
a
have wide application for treatment of carbohydrate mediated dis-
eases such as diabetes,1–3 cancer,4,5 HIV6 and certain forms of
hyperlipoproteinemia and obesity.7 Therefore, considerable
endeavors have been made to develop inhibitors that can probe
2. Results and discussion
In order to find a lead compound, 2–9 were designed and syn-
thesized with isosteviol as starting material (Scheme 1). Initial syn-
thetic efforts were focused on structural modifications at C-15 and
C-16 positions of isosteviol 1. Treatment of isosteviol obtained by
acid hydrolysis of stevioside with CH3CH2Br and KOH in DMSO
afforded the corresponding ethyl ester of isosteviol 2 in 96%
yield.20 Compounds 3 and 4 were obtained, respectively, in good
yields by reduction of 1 and 2 with NaBH4 in C2H5OH at 0 °C.21
The stereostructure of compound 4 was confirmed through X-ray
crystallographic analysis (Fig. 1). Treatment of 4 with acrylic acid
in CH2Cl2 in the presence of DCC and DMAP furnished 5 in 85%
yield.
Compounds 6 and 7 were stereoselectively synthesized via an
one pot Tollens’ reaction in good yield (95%, 90%, respectively).22
The products were characterized by HRMS, IR and NMR, and the
stereostructure of compound 6 was confirmed by X-ray crystallo-
graphic analysis (Fig. 2). The mechanism of the one pot Tollens’
reaction was proposed as shown in Scheme 2. In addition, com-
pound 9 could be obtained by selective oxidation of 7 with PCC
the structure and function of
a
-glycosidase.8,9 To date, various
types of inhibitors have also been designed based on the structures
that resemble the glycosyl cations in a transition state of hydroly-
sis by glucosidase.10
Isosteviol (ent-16-ketobeyeran-19-oic acid 1) is a tetracyclic
diterpenoid with a beyerane skeleton, obtained by acid hydrolysis
of stevioside.11,12 In recent years, isosteviol derivatives have at-
tracted scientific attention because of their remarkably broad spec-
trum of biological activities including antihypertension,13 anti-
inflammatory,14 glucocorticoid agonist,15 antiproliferation,16 anti-
tumor17 and inhibition of ent-kaurene synthase.18 Especially, Wang
and co-workers reported that isosteviol can decrease the blood
glucose concentration in Zucker diabetic fatty rats,19 which
prompted us to study isosteviol derivatives to develop new
cosidase inhibitors for the treatment of diabetes.
a-glu-
* Corresponding author. Tel./fax: +86 371 67767200.
E-mail address: jctao@zzu.edu.cn (J.-C. Tao).
0968-0896/$ - see front matter Ó 2009 Published by Elsevier Ltd.
doi:10.1016/j.bmc.2009.01.017

Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
1465
O
O
OH
OH
iii
COOH
COOEt
ii
COOEt
5
3
4
ii
viii
12
17
11
9
20
O
O
13
O
C
1
14
16
vi
i
D
2
A10
15
8
5 B
3
Br
4
7
19
6
18
COOEt
2
COOEt
8
COOH
iv
1
v
O
OH
OH
OH
OH
vii
v
OH
COOEt
9
COOEt
COOH
6
7
Figure 2. X-ray structure of compound 6.
Scheme 1. Reagents and conditions: (i) EtBr, DMSO, KOH, rt, 3 h, 96%; (ii) NaBH4,
C2H5OH, 0 °C, 1 h, 92–96%; (iii) DCC/DMAP, acrylic acid, CH2Cl2, rt, 12 h, 85%; (iv)
HCHO, NaOH, C2H5OH, 60 °C, 1 h, 95%; (v) HCHO, C2H5ONa, C2H5OH, 60 °C, 3 h, 90%;
(vi) EtBr, DMSO, KOH, 80 °C, 3 h, 96%; (vii) PCC, CH2Cl2, rt, 1 h, 82%; (viii) NaBH4,
C2H5OH, rt, 1 h, 96%.
With compound 6 in hand, some efforts were carried out for
functional group conversion at the carboxyl group in order to
probe the effect of different ester group on inhibition activity
against a-glycosidase. Therefore a series of ester derivatives were
synthesized from 1,3-diol 6 and p-toluenesulfonates in presence
of K2CO3 in CH3CN (Scheme 3). Without functional protection,
the reaction could be carried out smoothly. It should be noted that
the use of p-toluenesulfonates instead of alcohols could not only
improve the reaction rates, but also avoid the intermolecular con-
densation of compound 6. The structures of 10a–10j were charac-
terized by NMR, IR and HRMS spectra, respectively, and the
stereostructure of 10f was confirmed by X-ray crystallographic
analysis (Fig. 3).
in CH2Cl2, and treatment of 9 with HCHO in presence of C2H5ONa
in C2H5OH also gave the corresponding 7, which elucidated the
rationality of the proposed mechanism.
Meanwhile, treatment of 2 with an excess bromoethane in
DMSO in the presence of KOH under reflux gave 15-bromoisosteviol
ethyl ester 8, and the stereostructure of 15-bromoisosteviol ethyl
ester was confirmed according to X-ray crystallographic analysis
of 15-bromoisosteviol methyl ester.23 Unexpectedly, treatment of
8 with excess sodium borohydride in C2H5OH gave the debrominat-
Over the years, the literature has provided many examples of
ed compound instead of the expected a
-bromohydrin.24
a
,b-unsaturated ketones which possess interesting biological prop-
erties.25,26 It suggested that there would be potential interest in
simpler lipophilic structures containing an -methylene cyclo-
pentanone subunit. In this regard, an isosteviol derivative contain-
ing -methylene cyclopentanone fragment (13) was synthesized
The results obtained above showed that the newly introduced
hydroxymethyl or bromine group at C-15 was always stereoselec-
tively posited on exo position. From the crystal structure of com-
pounds 4 and 6, we found that the steric hindrance of C10-CH3
and ring C may be the reason for that substituent at C-15 could
not be posited on the endo position. Meanwhile, the newly intro-
duced hydroxy group at C-16 was stereoselectively posited on
the endo position because of the steric hindrance effects of C13-
CH3 and ring C.
a
a
from 7 as shown in Scheme 4. Selective esterification of 1,3-diol
7 with benzoyl chloride in presence of Et3N in toluene gave the
corresponding 1,3-diol monoester 11b (85%). Then, 1,3-diol mono-
esters 11b was oxidized by PDC to give thecorresponding production
12b, which was treated with 1,8-diazabicyclo[5.4.0]undec-7-ene
(DBU) in pyridine at 80 °C for 12 h to give the a-methylene ketone
13 in moderate yield (61%). The regioselectivity of 1,3-diol mono-
ester 11b was confirmed by X-ray crystallographic analysis of
In vitro activity screening of the above isosteviol derivatives
showed that 3–9 demonstrated
(Table 2), especially, compound 7 had higher inhibition activity
against -glycosidase (7 vs 6). Therefore, the introduction of hy-
droxyl, hydroxymethyl and ester group may enhance the inhibition
activity against -glycosidase.
a-glycosidase inhibition activity
a
12b (Fig. 4). In order to improve the yield of
a-methylene ketone
13, the reaction condition was optimized. Selective esterification
of 1,3-diol 7 with acetyl chloride instead of benzoyl chloride in
presence of Et3N gave the corresponding 1,3-diol monoester 11a
a
in good yield (96%), then
a-methylene ketone 13 was easily ob-
tained by oxidation and b-elimination in good yield.
Treatment of isosteviol 1 and its ethyl ester 2 with m-chloroper-
oxybenzoic acid (m-CPBA) in CH2Cl2 gave the corresponding
lactones 14a and 14b by Baeyer–Villiger oxidation.27,28 The struc-
ture of the product 14b was confirmed unambiguously by the dis-
appearance of the signals due to the carbonyl groups in the IR and
13C NMR spectra and the presence of lactone signal at dC 172. The
regioselectivity of 14b was confirmed by X-ray crystallographic
analysis (Fig. 5).
The Fischer indole reaction has remained an extremely impor-
tant and useful method for the synthesis of a variety of indole
intermediates and biologically active compounds.29–31 So, indole
isosteviol derivatives 15a and 15b were obtained using acetic acid
saturated with gaseous HCl as catalyst via Fischer reaction in good
Figure 1. X-ray structure of compound 4.

1466
Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
OH_ HCHO
+
H_ HCOOH
O
O
+
OH
OH
HCHO
OH_
OH
COOEt
2
COOEt
9
COOEt
7
Scheme 2. Proposed mechanism for synthesis of compound 7.
uct.35 In order to investigate the behavior of the 16-E-oxime iso-
steviol, further study was performed under different acid
OH
OH
a
conditions in Table 1. The results obtained showed that lactam
products accompanied with olefinic nitrile products occurred
simultaneously under different acid conditions. When H2SO4
(acetone, 40 °C) was used, beckmann fragmentation occurred read-
ily, giving rise to a 3:1 mixture of olefinic nitriles 20a and 20b,
which could not be separated by chromatography method, due to
the olefinie isomers had very similar physical properties and
polarities.
COOH
6
OH
OH
K2CO3
K2CO3
ROH + TsCl
TsOR
COOR
10
10a R=CH3
10f R=C(CH3)3
10b R=CH2CH2CH3
10c R=CH(CH3)2
10d R=CH2CH2CH2CH3
10e R=CH2CH(CH3)2
10g R=CH2CH2CH(CH3)2
10h R=CH2CH=CH2
10i R=CH2CH2OH
10j R=CH2C6H5
The inhibitory activities of these compounds were then evalu-
ated against
a-glucosidase as described in Table 2. Several inter-
esting structure–activity relationships have been obtained from
the analyses of the IC50 values of compounds 1–9. The first obser-
vation is that all the hydroxyl and hydroxymethyl substituted
isosteviols exhibit much higher inhibitory activities than the pre-
cursor isosteviol. This result is in agreement with the fact that
hydroxyl isosteviols always possess a variety of biological activi-
ties,36 and suggest that modification of isosteviol with hydroxyl
groups onto the beyerane skeleton is an efficient approach to in-
crease their inhibitory activities. It was also found that inhibitory
activities show significant dependence on the number of hydroxyl
groups. For example, compound 7 bearing two hydroxyl groups
has stronger activity than compound 4 bearing one hydroxyl
group. In addition, isosteviol derivatives containing carboxy group
show much lower bioactivity than their ester derivatives (3 vs 4
and 6 vs 7).
Scheme 3. Syntheses of 10.
yields (80%, 91%)32 In the 1H and 13C NMR spectra of 15b, addi-
tional resonances were observed at dH 7.05, 7.07, 7.32, 7.67, 7.81
and dC 111.6, 119.5, 119.6, 119.7, 121.8, 124.9, 138.8, 148.6, respec-
tively, suggesting the introduction of indole fragment.
Oximation of isosteviol 1 and its ethyl ester 2 with hydroxyl-
amine hydrochloride in presence of sodium bicarbonate in ethanol
gave the corresponding 16-E-oxime isosteviol 16a and 16-E-oxime
isosteviol ethyl ester 16b,27 which were further converted to the
16-amino isosteviol 17a and 16-amino isosteviol ethyl ester 17b,
respectively, by reduction of the oxime with Ni/H2 in THF in good
yields (82%, 88%). Unexpectedly, when ethanol was used instead of
THF in the reduction reaction of 16-E-oxime isosteviol ethyl ester
16b, 16-aminoethyl isosteviol ethyl ester 18 was obtained.
It is well known that certain oxime when subjected to Beck-
mann rearrangement condition33,34 do not rearrange into lactam
but undergo Beckmann fragmentation into olefinic nitrile prod-
The ester derivatives 10a–10j obtained from 1,3-diol 6 have
better inhibitory activities than their precursor isosteviol. But the
increase in bioactivity is not linear. The above results suggested
that the ester derivatives containing hydrophobic ester group exhi-
bit much higher inhibitory activities than that containing hydro-
philic ester group (10a, 10b, 10d vs 10i).
It was reported that compounds containing
tone show much high cytotoxic activities.25 However, isosteviol
derivative containing -methylene cyclopentanone fragment 13
show much lower inhibitory activity against -glucosidase. In
a,b-unsaturated ke-
a
a
addition, introduction of lactone, amine and oxime group into
the isosteviol structure result in higher inhibition activities ( 14,
16, 17, 18 vs 1), especially, the lactam derivative 19b (IC50
= 72.4
more inhibitory potency against
ring fused heterocyclic analogues might deserve some attention
for further -glucosidase inhibition activities design.
lM), and indole derivative 15b (IC50 = 68.2
lM) exhibit
a
-glucosidase, indicating that D-
a
3. Conclusion
In summary, a series of novel compounds containing hydroxyl,
hydroxymethyl group and heteroatom-containing frameworks
fused with isosteviol structure have been successfully synthesized
in high yields, and their inhibitory activities against
were evaluated. The results obtained revealed that these isosteviol
derivatives were capable of inhibiting -glucosidase with moder-
a-glucosidase
Figure 3. X-ray structure of compound 10f.
a

Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
1467
OH
OH
OH
O
iii
O
iv
ii
OCOR
OCOR
11a R=CH3
11b R=C6H5
COOEt
i
12a R=CH3
COOEt
COOEt
COOEt
COOEt
13
7
12b R=C6H5
NH
N
O
vii
x
CN
OH
O
+
COOR
ix
19a
COOR 16a R=H
R=H
COOR 1 R=H
20a+20b
16b
19b R=CH2CH3
R=CH2CH3
2 R=CH2CH3
v
vi
viii
O
O
NHEt
NH2
NH
COOR
COOEt
COOR
14a R=H
COOR
17a
R=H
17b R=CH2CH3
18
15a
14b
R=H
R=CH2CH3
15b R=CH2CH3
Scheme 4. Reagents and conditions: (i) HCHO, C2H5ONa, C2H5OH, 60 °C, 3 h, 90%; (ii) RCOCl/Et3N, toluene, rt, 2 h, (85–96%); (iii) PDC, CH2Cl2, rt, 3 h, (71–79%); (iv) DBU, pyr,
80 °C, 6–12 h, (61–85%); (v) m-CPBA, CH2Cl2, 0 °C, 5 h, (60–71%); (vi) HCl/AcOH, C6H5NHNH2, reflux, 3 h, (80–91%); (vii) HONH3Cl, NaHCO3, C2H5OH, 60 °C, 2 h, (90–95%); (viii)
Ni, H2, THF, 40 °C, 2 h, (82–88%); (ix) Ni, H2, C2H5OH, 40 °C, 2 h, 86%; (x) TsCl/DMF, 80 °C, or H2SO4/acetone, 40 °C, or BF3ÁOEt2/toluene, 100 °C.
ate to good activities, indicating that structural modification of iso-
steviol is a practical approach to increase their inhibitory activities.
Among all the derivatives, indole derivative 15b showed the high-
by Waters Q-Tof micro mass spectrometer. X-ray analysis was ta-
ken on a Rigaku RAXIS-IV.
est activities, and thus may be exploitable as potentially potent
glucosidase inhibitors. Further efforts aiming at developing potent
-glucosidase inhibitors based on appropriately modified D-ring
fused heterocyclic analogues are continuing in our laboratory,
which will be reported in due course.
a-
4.2. General procedure for a-glucosidase inhibition assay
a
Inhibition rate was determined at 37 °C in 0.067 M K2HPO4/
KH2PO4 buffer (pH 6.8). The reaction mixture contained 40 l of
enzyme solution, 40 l of inhibitor and 20 l of substrate. The sub-
strate and -glucosidase (Baker’s yeast) were purchased from Sig-
l
l
l
a
ma Chemical Co. (St. Louis, MO, USA). Both inhibitor and substrate
were first dissolved in dimethylsulfoxide (DMSO), and then diluted
with 0.067 M K2HPO4/KH2PO4 buffer to make the final concentra-
tion of DMSO 10%. The enzymatic reaction was started after incu-
bation of the enzyme (0.04 U/mL) for 30 min in the presence of the
inhibitor (0.1 mM) by the addition of substrate (0.5 mM). The mix-
ture was incubated at 37 °C for 5 min, and the reaction was
quenched by the addition of 0.1 M Na2CO3 (pH 9.8). The absorption
at 405 nm was measured immediately and taken as the relative
rate for the hydrolysis of substrate. All the experiment was carried
out in triplicate.
4. Experimental
4.1. General methods
All reagents and solvents were obtained from commercial sup-
pliers. All the reactions were monitored by TLC. Melting points
were determined on a Beijing Keyi XT5 apparatus and the temper-
ature was not corrected. IR spectra were recorded as KBr pellets on
a Thermo Nicolet (IR200) Spectrometer. 1H and 13C NMR spectra
were recorded on a Bruker DPX-400 spectrometer at 400 and
100 MHz with TMS as internal standard. Mass spectra were taken
Figure 4. X-ray structure of compound 12b.
Figure 5. X-ray structure of compound 14b.

1468
Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
Table 1
Beckmann rearrangement and fragmentation of 16-E-oxime isosteviol ethyl ester 16b
NH
N
OH
acid catalyst
CN
CN
O
+
+
COOEt
19b
COOEt
20a
COOEt
20b
COOEt
16b
Entry
Catalyst/solvent
T (°C)
Time (h)
Yield of 19b (%)a
Yield of 20a + 20b (%)a
1
2
3
TsCl/DMF
H2SO4/acetone
BF3ÁOEt2/toluene
80
40
100
10
6
12
55
6
77
38
88 (20a/20b = 3:1)b
22
a
Yields of isolated products.
b
Determined by 1H NMR spectra.
4.3. The preparation of isosteviol derivatives
with MgSO4 and concentrated under vacuum to give white powder
3 (0.294 g, 92%), mp 168–169 °C; IR (KBr): 3475, 2990, 2943, 2896,
4.3.1. ent-16-Oxobeyeran-19-oic acid (1)12
2841, 1653, 1453, 1371, 1187, 1056, 998, 621 cmÀ1 1H NMR
;
Isosteviol 1 was synthesized by hydrolysis of stevioside with di-
lute sulfuric acid. Mp 228–230 °C; IR (KBr): 3455, 2954, 2927,
2852, 2679, 1738, 1697, 1474, 1453, 1406, 1372, 1320, 1271,
(400 MHz, CDCl3): d 3.64 (m, 1H), 2.01 (d, J = 12.8 Hz, 1H), 1.76–
1.62 (m, 5H), 1.59–1.52 (m, 3H), 1.45–1.41 (m, 2H), 1.31–1.17
(m, 3H), 1.09 (s, 3H), 1.06–0.86 (m, 6H), 0.82 (s, 3H), 0.75 (s, 3H);
HRMS (ESI) calcd for C20H33O3 [M+H]+ 321.2430. Found: 321.2425.
1238, 1179, 950, 797 cmÀ1 1H NMR (400 MHz, CDCl3): d 2.64
;
(dd, J = 18.6, 3.8 Hz, 1H), 2.17 (d, J = 13.4 Hz, 1H), 1.90–1.82 (m,
2H), 1.81 (d, J = 18.6 Hz 1H), 1.77–1.38 (m, 10H), 1.26 (s, 3H),
1.22–1.14 (m, 3H), 1.07–0.99 (m, 1H), 0.98 (s, 3H), 0.95–0.88 (m,
1H), 0.79 (s, 3H); HRMS (ESI, m/z) calcd for C20H30O3Na [M+Na]+
341.2093. Found: 341.2085.
4.3.4. Ethyl ent-16b-hydroxybeyeran-19-oate (4)
A solution of isosteviol ethyl ester 2 (0.346 g, 1 mmol) and so-
dium borohydride (0.057 g, 1.5 mmol) in dry ethanol (10 mL) was
stirred at 0 °C for 1 h. After that the reaction mixture was concen-
trated under vacuum, and extracted with CHCl3 and H2O. At last
the organic layer was washed with saturated NaCl aqueous solu-
tion, dried with MgSO4 and concentrated under vacuum to give
white powder 4 (0.334 g, 96%), mp 152–153 °C; IR (KBr): 3533,
2978, 2939, 2880, 2837, 1700, 1460, 1374, 1318, 1231, 1178,
4.3.2. Ethyl ent-16-oxobeyeran-19-oate (2)
The isosteviol ethyl ester 2 was obtained by treating isosteviol 1
with CH3CH2Br and KOH in DMSO at room temperature in good
yield (92%) according to the literature method.20 Mp 125–127 °C;
IR (KBr): 2957, 2926, 2847, 1726, 1451, 1377, 1227, 1146, 1096,
1151, 1049 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.09 (q, J = 7.2 Hz,
;
1024 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.10 (q, J = 7.2 Hz, 2H),
;
2H), 3.85 (q, J = 4.8 Hz, 1H), 2.16 (d, J = 13.2 Hz, 1H), 1.81–1.51
(m, 11H), 1.26 (t, J = 7.2 Hz, 3H), 1.23–1.18 (m, 1H), 1.16 (s, 3H),
1.04–0.93 (m, 4H), 0.90 (s, 3H), 0.88–0.86 (m, 1H), 0.74 (s, 3H);
13C NMR (100 MHz, CDCl3): d 177.6, 80.6, 59.9, 57.1, 55.8, 55.2,
43.7, 42.8, 42.0, 41.7, 39.9, 38.1, 38.0, 33.7, 29.0, 24.9, 21.7, 20.4,
18.9, 14.1, 13.3; HRMS (ESI, m/z) calcd for C21H34O4Na [M+Na]+
371.2562. Found: 371.2554.
2.96 (dd, J = 18.4, 2.8 Hz, 1H), 2.17 (d, J = 11.2 Hz, 1H), 1.99 (d,
J = 18.8 Hz, 1H), 1.88–1.58 (m, 8H), 1.47–1.28 (m, 6H), 1.26 (t,
J = 7.2 Hz, 3H), 1.24–1.19 (m, 2H), 1.18 (s, 3H), 1.07 (s, 3H), 1.05–
0.84 (m, 1H), 0.77 (s, 3H); HRMS (ESI, m/z) calcd for C22H34O3Na
[M+Na]+ 369.2406. Found: 369.2400.
4.3.3. ent-16b-Hydroxybeyeran-19-oic acid (3)21
A solution of isosteviol 1 (0.318 g, 1 mmol) and sodium borohy-
dride (0.057 g, 1.5 mmol) in dry ethanol (10 mL) was stirred at 0 °C
for 1 h. After that the reaction mixture was concentrated under
vacuum, and extracted with CHCl3 and H2O. At last the organic
layer was washed with saturated NaCl aqueous solution, dried
4.3.5. Ethyl ent-16b-acryloxybeyeran-19-oate (5)
A mixture of the compound 4 (0.348 g, 1 mmol), acrylic acid
(0.792 g, 1.1 mmol), DCC (0.412 g, 2 mmol) and DMAP (0.024 g,
0.2 mmol) was stirred at room temperature. After stirring for 12 h,
the reaction mixture was filtered, and the filtrate was concentrated.
Table 2
Inhibition activities of isosteviol derivatives against
a-glucosidase
Compound
a
-Glucosidaseb
Compound
a
-Glucosidaseb
Compound
a
-Glucosidaseb
1
2
3
4
5
6
7
8
>200
>200
156.3
132.1
>200
148.6
86.2
10d
10e
10f
10g
10h
10i
87.2
14a
14b
15a
15b
16a
16b
17a
17b
18
138.6
118.4
83.2
68.2
92.1
88.9
>200
91.2
113.6
81.6
72.4
NIa
102.5
113.8
115.6
143.2
>200
>200
132.5
112.4
>200
>200
>200
10j
NIa
11a
11b
12a
12b
13
9
143.2
96.5
85.4
10a
10b
10c
19a
19b
20
97.2
a
No inhibition at 200 lV.
IC50 (lM).
b

Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
1469
The residue was purified by column chromatography on silica
(petroleum ether/ethyl acetate = 6:1, v/v) to give product
(0.341 g, 85%). IR (KBr): 3101, 2950, 2847, 1723, 1625, 1455,
1405, 1378, 1194, 1151, 1060, 981, 811 cmÀ1 1H NMR (400 MHz,
CDCl3): d 6.38 (d, J = 17.2 Hz, 1H), 6.13 (dd, J = 17.2, 10.4 Hz, 1H),
5.81 (d, J = 10.4 Hz, 1H), 4.80 (q, J = 4.8 Hz, 1H), 4.07 (m, 2H), 2.17
(d, J = 13.6 Hz, 1H), 1.92–1.68 (m, 7H), 1.61–1.33 (m, 7H), 1.25 (t,
J = 7.2 Hz, 3H), 1.15 (s, 3H), 1.09–0.94 (m, 4H), 0.90 (s, 3H), 0.87–
0.84 (m, 1H), 0.70 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.5,
166.4, 130.2, 128.9, 81.7, 59.9, 57.0, 55.7, 54.8, 43.7, 42.4, 41.6,
41.5, 40.6, 39.9, 38.5, 38.0, 34.6, 28.9, 24.9, 21.7, 20.2, 18.9, 14.1,
13.2; HRMS (ESI, m/z) calcd for C25H39O4 [M+H]+ 403.2848. Found:
403.2835.
3H), 1.18–1.15 (m, 1H), 1.09 (s, 3H), 1.04–0.87 (m, 3H), 0.75 (s,
3H); 13C NMR (100 MHz, CDCl3): d 216.5, 177.8, 60.8, 57.9, 56.8,
56.6, 50.7, 49.0, 44.3, 43.7, 40.1, 39.4, 39.0, 38.5, 38.4, 29.5, 21.5,
21.4, 20.6, 19.6, 14.8, 14.3; HRMS (ESI, m/z) calcd for C22H34NBrO3
[M+H]+ 425.1691. Found: 425.1681.
5
;
4.3.9. Ethyl ent-15a-hydroxymethyl-16-oxobeyeran-19-oate (9)
A mixture of the compound 7 (0.378 g, 1 mmol) and PCC
(0.236 g, 1.1 mmol) was stirred at room temperature for 1 h. Then
the reaction mixture was filtered, and the filtrate was concen-
trated. The residue was purified by column chromatography on sil-
ica (petroleum ether/ethyl acetate = 7:1, v/v) to give product 9
(0.308 g, 82%). Mp 155–157 °C; IR (KBr): 3534, 2958, 2857, 1735,
1721, 1462, 1151 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.10 (m,
;
4.3.6. ent-15a-Hydroxymethyl-16b-hydroxybeyeran-19-oic acid
2H), 3.90 (m, 1H), 3.70 (t, J = 10.4 Hz, 1H), 2.52 (m, 1H), 2.50 (m,
1H), 2.19 (d, J = 13.3 Hz, 1H), 1.89–1.69 (m, 8H), 1.42–1.29 (m,
4H), 1.27 (t, J = 7.2 Hz, 3H), 1.19 (s, 3H), 1.18–1.10 (m, 2H), 0.98
(s, 3H), 0.97–0.80 (m, 2H), 0.75 (s, 3H); 13C NMR (100 MHz, CDCl3):
d 226.1, 177.2, 60.4, 60.1, 57.1, 56.7, 52.9, 52.5, 48.4, 43.6, 40.5,
39.6, 38.2, 37.8, 37.0, 35.2, 28.9, 21.6, 19.8, 19.6, 18.8, 14.1, 13.3;
HRMS (ESI, m/z) calcd for C23H36O4Na [M+Na]+ 399.2511. Found:
399.2514.
(6)22
To a stirred solution of compound 1 (0.318 g, 1 mmol) and
NaOH (0.08 g, 2 mmol) in ethanol (20 mL) was added 37% formal-
dehyde aqueous solution (2 mL). After stirring for 1 h at 60 °C, the
mixture was concentrated under vacuum, and extracted with
CHCl3 and H2O, at last the organic layer was washed with saturated
NaCl aqueous solution, dried with MgSO4 and concentrated under
vacuum to give white powder 6 (0.332 g, 95%), mp 233–235 °C; IR
(KBr): 3462, 2945, 2927, 2846, 1696, 1456, 1072, 1052 cmÀ1 1H
;
4.3.10. General procedure for synthesis of compounds 10a–10j
A mixture of various alcohols (0.032–0.108 g, 1 mmol) and 4-
methylphenylsulfonyl chloride (0.216 g, 1 mmol) in dry acetoni-
trile were stirred at 50 °C in the presence of K2CO3 (0.414 g,
3 mmol). After completion of the reaction monitored by TLC, com-
pound 6 was added. After stirring of the mixture overnight, the
mixture was concentrated under vacuum, and extracted with
CHCl3 and H2O. At last the organic layer was washed with satu-
rated NaCl aqueous solution, dried with MgSO4 and concentrated
under vacuum. The residue was purified by column chromatogra-
phy on silica to give product 10.
NMR (400 MHz, acetone-d6): d 3.83 (dd, J = 10.4, 5.2 Hz, 1H), 3.62
(d, J = 4.8 Hz, 1H), 3.50 (t, J = 9.6 Hz, 1H), 3.30 (s, 2H), 2.12–1.99
(m, 2H), 1.95–1.70 (m, 6H), 1.56–1.51 (m, 1H), 1.44–1.34 (m,
2H), 1.17 (s, 3H), 1.15–0.90 (m, 6H), 0.88 (s, 3H), 0.87 (s, 3H); 13C
NMR (100 MHz, acetone-d6): d 179.0, 82.3, 62.4, 57.8, 56.7, 54.5,
50.2, 43.1, 42.6, 40.5, 39.1, 38.2, 38.0, 34.9, 33.8, 29.1, 25.6, 22.3,
19.4, 19.0, 13.4; HRMS (ESI, m/z) calcd for C21H34O4Na [M+Na]+
373.2355. Found: 373.2358.
4.3.7. Ethyl ent-15a-hydroxymethyl-16b-hydroxybeyeran-19-
oate (7)22
To a stirred solution of compound 2 (0.346 g, 1 mmol) and
C2H5ONa (0.136 g, 2 mmol) in ethanol (20 mL) was added 37%
formaldehyde aqueous solution (2 mL). After stirring for 3 h at
60 °C, the mixture was concentrated under vacuum, and extracted
with CHCl3 and H2O, at last the organic layer was washed with sat-
urated NaCl aqueous solution, dried with MgSO4 and concentrated
under vacuum to give white powder 7 (0.34 g, 90%). Mp 181–
182 °C; IR (KBr): 3435, 2940, 2838, 1720, 1458, 1378, 1234,
4.3.10.1. Methyl ent-15a-hydroxymethyl-16b-hydroxybeyeran-
19-oate (10a). Yield 93%; mp 181–182 °C; IR (KBr): 3416, 2948,
2845, 1720, 1452, 1370, 1327, 1235, 1187, 1154, 1096,
1070 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.00 (dd, J = 9.9, 5.2 Hz,
;
1H), 3.63 (s, 4H), 3.48 (t, J = 10.3 Hz, 1H), 2.16 (d, J = 13.2 Hz, 1H),
2.03–1.91 (m, 5H), 1.82–1.56 (m, 6H), 1.43–1.37 (m, 2H), 1.26–
1.17 (m, 1H), 1.16 (s, 3H), 1.08–0.97 (m, 3H), 0.94 (s, 3H), 0.92–
0.86 (m, 1H), 0.75 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.9,
86.8, 64.9, 57.5, 57.0, 54.1, 51.2, 50.2, 43.6, 42.4, 40.8, 39.5, 38.1,
37.8, 34.7, 33.0, 28.8, 25.0, 22.1, 19.4, 18.8, 12.9; HRMS (ESI, m/z)
calcd for C22H36O4Na [M+Na]+ 387.2511. Found: 387.2514.
1179, 1153, 1123 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.09 (q,
;
J = 7.2 Hz, 2H), 3.98 (dd, J = 9.7, 5.0 Hz, 1H), 3.63 (d, J = 4.7 Hz,
1H), 3.56 (t, J = 10.2 Hz, 1H), 2.16 (d, J = 13.0 Hz, 1H), 2.05 (m,
1H), 1.83–1.56 (m, 9H), 1.43–1.37 (m, 2H), 1.26 (t, J = 7.2 Hz, 3H),
1.22–1.19 (m, 1H), 1.16 (s, 3H), 1.08–0.95 (m, 4H), 0.94 (s, 3H),
0.88–0.86 (m, 1H), 0.78 (s, 3H); 13C NMR (100 MHz, CDCl3): d
177.4, 86.7, 64.9, 60.0, 57.5, 57.0, 54.2, 50.2, 43.6, 42.4, 40.8, 39.6,
38.1, 37.9, 34.8, 33.0, 28.9, 25.0, 22.1, 19.5, 18.8, 14.1, 13.2; HRMS
(ESI, m/z) calcd for C23H38O4Na [M+Na]+ 401.2668. Found:
401.2664.
4.3.10.2. n-Propyl ent-15a-hydroxymethyl-16b-hydroxybeyer-
an-19-oate (10b). Yield 90%; mp 120–121 °C; IR (KBr): 3405,
2954, 2847, 1723, 1468, 1454, 1387, 1374, 1232, 1180, 1123,
1072, 1052, 1005, 951, 722, 611 cmÀ1 1H NMR (400 MHz, CDCl3):
;
d 3.98 (m, 3H), 3.63 (d, J = 4.7 Hz, 1H), 3.49 (t, J = 10.4 Hz, 1H),
2.14–2.01 (m, 4H), 1.90–1.50 (m, 9H), 1.40–1.36 (m, 2H), 1.26–
1.17 (m, 1H), 1.16 (s, 3H), 1.10–0.98 (m, 4H), 0.97 (t, J = 7.4 Hz,
3H), 0.94 (s, 3H), 0.92–0.86 (m, 1H), 0.77 (s, 3H); 13C NMR
(100 MHz, CDCl3): d 177.5, 86.7, 65.8, 64.9, 57.5, 57.0, 54.1, 50.2,
43.7, 42.4, 40.8, 39.5, 38.1, 37.9, 34.7, 33.0, 28.9, 25.0, 22.1, 21.8,
19.4, 18.8, 13.1, 10.7; HRMS (ESI, m/z) calcd for C24H40O4Na
[M+Na]+ 415.2824. Found: 415.2831.
4.3.8. Ethyl ent-15a-bromo-16-oxobeyeran-19-oate (8)
To a stirred solution of compound 2 (0.346 g, 1 mmol) and KOH
(0.112 g, 2 mmol) in DMSO (10 mL) was added CH3CH2Br (2 mL),
and the mixture was heated at 80 °C for 3 h. Then the mixture
was poured into H2O and extracted with CHCl3. The extract was
washed with water, dried and evaporated. The residue was purified
by column chromatography on silica (petroleum ether/ethyl ace-
tate = 9:1, v/v) to give product 8 (0.407 g, 96%). Mp 150–151 °C;
IR (KBr): 2956, 2934, 2873, 2851, 1747, 1716, 1454, 1378, 1239,
4.3.10.3. i-Propyl ent-15
an-19-oate (10c). Yield 92%; mp 137–138 °C; IR (KBr): 3440,
2938, 2848, 1719, 1455, 1375, 1234, 1179, 1153, 1109 cmÀ1 1H
a-hydroxymethyl-16b-hydroxybeyer-
1151, 1105, 1028 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.50 (d,
;
J = 2.4 Hz, 1H), 4.11 (q, J = 7.2 Hz, 2H), 2.24–2.17 (m, 2H), 1.93–
1.66 (m, 7H), 1.51–1.31 (m, 5H), 1.27 (t, J = 3.2 Hz, 3H), 1.19 (s,
;
NMR (400 MHz, CDCl3): d 4.97 (m, 1H), 3.99 (dd, J = 9.9, 5.2 Hz,
1H), 3.62 (d, J = 4.6 Hz, 1H), 3.50 (t, J = 10.3 Hz, 1H), 2.15 (d,

1470
Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
J = 13.0 Hz, 1H), 2.00 (m, 1H), 1.82–1.56 (m, 8H), 1.43–1.37 (m, 2H),
1.26 (d, J = 6.2 Hz, 3H), 1.23 (d, J = 6.2 Hz, 3H), 1.13 (s, 3H), 1.09–
0.96 (m, 6H), 0.94 (s, 3H), 0.90–0.82 (m, 1H), 0.80 (s, 3H); 13C
NMR (100 MHz, CDCl3): d 176.8, 86.7, 67.1, 64.9, 57.5, 56.9, 54.2,
50.2, 43.6, 42.4, 40.8, 39.6, 38.2, 37.9, 34.8, 33.0, 28.9, 25.0, 22.2,
21.7, 21.6,19.4, 18.8, 13.4; HRMS (ESI, m/z) calcd for C24H40O4Na
[M+Na]+ m/z 415.2824. Found: 415.2827.
2.17 (d, J = 13.2 Hz, 1H), 2.01 (m, 2H), 1.90–1.77 (m, 4H), 1.76–
1.55 (m, 3H), 1.43–1.20 (m, 3H), 1.18 (s, 3H) ,1.09–0.98 (m, 5H),
0.96 (s, 3H), 0.90–0.83 (m, 1H), 0.76 (s, 3H); 13C NMR (100 MHz,
CDCl3): d 177.1, 132.3 ,118.0, 86.6, 64.9, 64.9, 57.5, 57.1, 54.1,
50.2, 43.7, 42.5, 40.8, 39.5, 38.2, 37.9, 34.7, 33.1, 28.9, 25.0, 22.2,
19.5, 18.8, 13.2; HRMS (ESI, m/z) calcd for C24H38O4Na [M+Na]+
413.2668. Found: 413.2663.
4.3.10.4. n-Butyl ent-15
a
-hydroxymethyl-16b-hydroxybeyeran-
4.3.10.9. Hydroxyethyl ent-15a-hydroxymethyl-16b-hydroxybey-
19-oate (10d). Yield 91%; mp 99–100 °C; IR (KBr): 3423, 2944,
eran-19-oate (10i). Yield 93%; mp 176–177 °C; IR (KBr): 3411,
2873, 2847, 1721, 1458, 1373, 1323, 1230, 1178, 1151, 1125,
2942, 2847, 1720, 1457, 1386, 1323, 1231, 1178, 1153, 1072,
1096, 1068, 1052, 983 cmÀ1
;
1H NMR (400 MHz, CDCl3): d 4.00
1052 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.27 (m, 1H), 4.04 (m,
;
(m, 3H), 3.63 (m, 1H), 3.48 (t, J = 10.3 Hz, 1H), 2.16 (d, J = 13.2 Hz,
1H), 1.82–1.56 (m, 10H), 1.44–1.37 (m, 4H), 1.21–1.17 (m, 2H),
1.16 (s, 3H), 1.08–0.93 (m, 8H), 0.94 (s, 3H), 0.92–0.80 (m, 1H),
0.77 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.6, 86.7, 65.0, 64.0,
57.5, 57.0, 54.2, 50.2, 43.7, 42.4, 40.8, 39.6, 38.1, 37.9, 34.8, 33.1,
30.5, 29.0, 25.0, 22.2, 19.5, 19.4, 18.9, 13.7, 13.1; HRMS (ESI, m/z)
calcd for C25H42O4Na [M+Na]+ 429.2981. Found: 429.2986.
1H), 3.98 (dd, J = 9.9, 5.2 Hz, 1H), 3.82 (m, 2H), 3.54 (d, J = 4.3 Hz,
1H), 3.43 (t, J = 10.3 Hz, 1H), 2.16 (d, J = 13.2 Hz, 1H), 2.0 (m, 1H),
1.82–1.65 (m, 5H), 1.50–1.56 (m, 3H), 1.45–1.34 (m, 2H), 1.30–1.20
(m, 1H), 1.18 (s, 3H), 1.11–0.96 (m, 5H), 0.92 (s, 3H), 0.90–0.83 (m,
1H), 0.82 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.4, 85.5, 65.3,
64.0, 59.7, 57.3, 56.7, 53.9, 50.2, 43.4, 42.3, 40.4, 38.2, 37.9, 37.7,
34.5, 33.1, 28.6, 25.0, 21.9, 19.2, 18.6, 12.9; HRMS (ESI, m/z) calcd
for C23H38O5Na [M+Na]+ 417.2617. Found: 417.2614.
4.3.10.5. i-Butyl ent-15a-hydroxymethyl-16b-hydroxybeyeran-
19-oate (10e). Yield 91%; mp 123–124 °C; IR (KBr): 3407, 2951,
4.3.10.10. Benzyl ent-15a-hydroxymethyl-16b-hydroxybeyeran-
2876, 2848, 1722, 1466, 1369, 1327, 1180, 1152, 1126, 1072,
19-oate (10j). Yield 89%; mp 75–76 °C; IR (KBr): 3416, 3064, 3031,
1007 cmÀ1
;
1H NMR (400 MHz, CDCl3): d 3.99 (dd, J = 9.9, 5.2 Hz,
2936, 2847, 1723, 1656, 1626, 1499, 1455, 1369, 1328, 1230, 1175,
1H); 3.77 (m, 2H), 3.63 (d, J = 4.6 Hz, 1H), 3.49 (t, J = 10.3 Hz, 1H),
2.17 (d, J = 13.0 Hz, 1H), 2.05 (m, 2H), 1.93–1.60 (m, 12H), 1.40–
1.18 (m, 3H), 1.17 (s, 3H), 1.08–0.99 (m, 2H), 0.97 (d, J = 1.6 Hz,
3H), 0.95 (d, J = 1.7 Hz, 3H), 0.94 (s, 3H), 0.91–0.83 (m, 1H), 0.77
(s, 3H); 13C NMR (100 MHz, CDCl3): d 177.6, 86.7, 70.5, 65.0, 57.5,
57.0, 54.2, 50.3, 43.8, 42.5, 40.9, 39.6, 38.2, 37.9, 34.8, 33.1, 29.0,
27.6, 25.0, 22.2, 19.5, 19.4, 19.3, 18.9, 13.1; HRMS (ESI, m/z) calcd
for C25H42O4Na [M+Na]+ 429.2981. Found: 429.2981.
1147, 1123, 1096, 1050, 1008, 751, 734, 697 cmÀ1 1H NMR
;
(400 MHz, CDCl3): d 7.35 (m, 5H), 5.12 (d, J = 12.3 Hz, 1H), 5.01 (d,
J = 12.3 Hz, 1H), 3.83 (dd, J = 9.8, 5.0 Hz, 1H), 3.61 (d, J = 4.7 Hz,
1H), 3.40 (t, J = 10.2 Hz, 1H), 2.19 (d, J = 13.4 Hz, 1H), 1.93 (m, 1H),
1.90–1.49 (m, 8H), 1.42–1.34 (m, 2H), 1.22–1.13 (m, 1H), 1.18 (s,
3H), 1.08–0.95 (m, 4H), 0.93 (s, 3H), 0.92–0.82 (m, 2H), 0.68 (s,
3H); 13C NMR (100 MHz, CDCl3): d 177.2, 135.9, 128.4, 128.4,
128.2, 128.0, 128.0, 86.7, 66.1, 64.8, 57.5, 57.0, 54.1, 50.2, 43.7,
42.4, 40.8, 39.5, 38.1, 37.9, 34.7, 33.0, 28.8, 25.0, 22.2, 19.4, 18.8,
13.1; HRMS (ESI, m/z) calcd for C28H40O4Na [M+Na]+ 463.2824.
Found: 463.2820.
4.3.10.6. t-Butyl ent-15a-hydroxymethyl-16b-hydroxybeyeran-
19-oate (10f). Yield 94%; mp 156–157 °C; IR (KBr): 3431, 2940,
2848, 1721, 1685, 1458, 1391, 1368, 1323, 1254, 1148, 1126,
1054, 856 cmÀ1
;
1H NMR (400 MHz, CDCl3): d 3.99 (dd, J = 9.9,
4.3.11. General procedure for synthesis of compounds 11a and
11b
5.2 Hz, 1H), 3.62 (d, J = 4.6 Hz, 1H), 3.48 (t, J = 10.3 Hz, 1H), 2.10
(d, J = 13.0 Hz, 1H), 2.05 (m, 1H), 1.80–1.69 (m, 5H), 1.68–1.53
(m, 3H), 1.43 (s, 9H), 1.42–1.35 (m, 2H), 1.3–1.13 (m, 1H), 1.12
(s, 3H), 1.09–0.95 (m, 4H), 0.94 (s, 3H), 0.92–0.86 (m, 2H), 0.85
(s, 3H); 13C NMR (100 MHz, CDCl3): d 176.6, 86.7, 79.8, 65.0, 57.6,
56.9, 54.2, 50.2, 44.3, 42.6, 40.9, 39.8, 38.4, 38.2, 35.0, 33.2, 29.1,
28.0, 28.0, 28.0, 25.1, 22.3, 19.5, 19.0, 13.8; HRMS (ESI, m/z) calcd
for C25H42O4Na [M+Na]+ 429.2981. Found: 429.2983.
A mixture of compound 7 (0.378 g, 1 mmol) and Et3N (0.202 g,
1.1 mmol) in toluene (20 mL) was stirred at 0 °C for 10 min, and
RCOCl (1.1 mmol) in toluene (5 mL) was added dropwise to the
solution during 1 h, then the mixture was stirred at room temper-
ature for 1 h. It was poured into water and extracted with CHCl3.
The extract was washed with water, dried and evaporated. The res-
idue was purified by column chromatography on silica to give
product 11.
4.3.10.7. i-Pentyl ent-15a-hydroxymethyl-16b-hydroxybeyeran-
19-oate (10g). Yield 89%; mp 73–74 °C; IR (KBr): 3418, 2953,
4.3.11.1. Ethyl ent-15a-acetoxymethyl-16b-hydroxybeyeran-19-
2872 2847, 1721, 1463, 1371, 1324, 1232, 1177, 1151, 1052,
oate (11a). Yield 96%; mp 90–91 °C; IR (KBr): 3445, 2945, 2849,
1007, 963 cmÀ1
;
1H NMR (400 MHz, CDCl3): d 4.05 (m, 2H); 3.98
1740, 1721, 1458, 1384, 1368, 1238, 1181, 1151, 1127, 1051,
(d, J = 4.8 Hz, 1H), 3.63 (d, J = 4.8 Hz, 1H), 3.49 (t, J = 10.4 Hz, 1H),
2.15 (d, J = 15.2 Hz, 1H), 2.03 (m, 1H), 1.86–1.70 (m, 5H), 1.67–
1.64 (m, 4H), 1.63–1.49 (m, 4H), 1.42–1.18 (m, 3H), 1.15 (s, 3H),
1.09–0.97 (m, 3H), 0.96 (s, 3H), 0.92 (d, J = 1.8 Hz, 6H), 0.90–0.83
(m, 1H), 0.77 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.5, 86.7,
64.9, 62.7, 57.5, 57.0, 54.2, 50.2, 43.7, 42.4, 40.8, 39.6, 38.1, 37.9,
37.2 ,34.8, 33.1, 28.9, 25.2, 25.0, 22.4, 22.4, 22.1, 19.5, 18.9, 13.2;
HRMS (ESI, m/z) calcd for C26H44O4Na [M+Na]+ 443.3137. Found:
443.3139.
1031, 977 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.34 (dd, J = 10.6,
;
5.4 Hz, 1H), 4.09 (q, J = 7.1 Hz, 2H), 3.95 (t, J = 10.3 Hz, 1H), 3.49
(d, J = 4.7 Hz, 1H), 2.19–2.11 (m, 2H), 2.09 (s, 3H), 1.90–1.61 (m,
10H), 1.42–1.39 (m, 2H), 1.25 (t, J = 7.1 Hz, 3H), 1.22–0.93 (m, 6H),
0.92 (s, 3H), 0.90–0.77 (m, 2H), 0.75 (s, 3H); 13C NMR (100 MHz,
CDCl3): d 177.3, 171.2, 86.0, 66.8, 60.0, 57.5, 57.0, 54.0, 47.2, 43.6,
42.7, 40.9, 39.6, 38.1, 37.9, 34.8, 33.1, 28.9, 25.0, 22.0, 21.2, 19.4,
18.8, 14.1, 13.1; HRMS (ESI, m/z) calcd for C25H40O5Na [M+Na]+
443.2773. Found: 443.2774.
4.3.10.8. Propenyl ent-15
a
-hydroxymethyl-16b-hydroxybeyer-
4.3.11.2. Ethyl ent-15
19-oate (11b). Yield 85%; mp 115–117 °C; IR (KBr): d 3427, 2926,
2882, 1719, 1601, 1456, 1275, 1152, 1119 cmÀ1 1H NMR
(400 MHz, CDCl3): 8.07 (d, J = 8.0 Hz, 2H), 7.58 (t, J = 8.0 Hz, 1H),
7.46 (t, J = 7.8 Hz, 2H), 4.60 (dd, J = 10.6, 5.4 Hz, 1H), 4.24 (t,
J = 10.4 Hz, 1H), 4.09 (q, J = 7.2 Hz, 2H), 3.67 (d, J = 4.8 Hz, 1H),
a-benzoyloxymethyl-16b-hydroxybeyeran-
an-19-oate (10h). Yield 96%; mp 66–67 °C; IR (KBr): 3426, 3084,
2943, 2848, 1722, 1646, 1458, 1380, 1323, 1229, 1175, 1149, 1126,
1053, 981 cmÀ1; 1H NMR (400 MHz, CDCl3): d 5.92 (m, 1H), 5.33 (d,
J = 17.2 Hz, 1H), 5.23 (d, J = 10.4 Hz, 1H), 4.53 (m, 2H), 3.98 (q,
J = 4.8 Hz, 1H), 3.63 (d, J = 4.8 Hz, 1H), 3.48 (t, J = 10.4 Hz, 1H),
;

Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
1471
2.33 (m, 1H), 2.17 (d, J = 13.6 Hz, 1H), 1.87–1.40 (m, 10H), 1.25 (t,
J = 7.2 Hz, 3H), 1.17 (s, 3H), 1.15–0.99 (m, 4H), 0.94 (s, 3H), 0.92–
0.83 (m, 3H), 0.80 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.4,
170.7, 131.6, 129.7, 129.3, 129.3, 129.0, 129.0, 87.1, 66.8, 61.7,
58.7, 56.2, 56.0, 50.9, 47.6, 44.2, 40.3, 39.5, 38.2, 37.1, 36.2, 34.5,
28.8, 25.4, 20.0, 19.2, 18.3, 14.1, 13.1; HRMS (ESI, m/z) calcd for
C30H42O5Na [M+Na]+ 505.2930. Found: 505.2931.
and the filtrate was concentrated. The residue was purified by col-
umn chromatography on silica to give product 14.
4.3.14.1. Lactone of isosteviol (14a)27
271 °C; IR (KBr): 3438, 2949, 2925, 2847, 1721, 1693, 1454,
1372, 1243, 1156, 771 cmÀ1 1H NMR (400 MHz, CDCl3): d 3.14
. Yield 60%; mp 270–
;
(dd, J = 18.6, 2.5 Hz, 1H), 2.18 (d, J = 13.4 Hz, 1H), 2.07–1.79 (m,
5H), 1.71 (br d, J = 13.4 Hz, 1H), 1.61–1.37 (m, 7H), 1.34 (s, 3H),
1.25 (s, 3H), 1.23–0.97 (m, 5H), 0.87 (s, 3H); HRMS (ESI, m/z) calcd
for C20H30O4Na [M+Na]+ 357.2042. Found: 357.2068.
4.3.12. General procedure for synthesis of compounds 12a–12b
A mixture of the compound 11 (1 mmol) and PDC (0.412 g,
2 mmol) in CH2Cl2 was stirred at room temperature for 3 h, then
the reaction mixture was filtered, and the filtrate was concen-
trated. The residue was purified by column chromatography on sil-
ica to give product 12.
4.3.14.2. Lactone of isosteviol ethyl ester (14b). Yield 71%; mp
153–154 °C; IR (KBr): 2958, 2925, 1715, 1474, 1446, 1377, 1239,
1146, 1020, 977 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.09 (m, 2H),
;
3.10 (dd, J = 18.8, 2.6 Hz, 1H), 2.17 (d, J = 13.2 Hz, 1H), 2. 04 (d,
J = 18.8 Hz, 1H), 1.99–1.54 (m, 9H), 1.46–1.37 (m, 3H), 1.34 (s,
3H), 1.26 (t, J = 7.2 Hz, 3H), 1.17 (s, 3H), 1.09–0.82 (m, 5H), 0.78
(s, 3H); 13C NMR (100 MHz, CDCl3): d 177.0, 172.7, 80.3, 60.1,
57.2, 55.8, 47.7, 43.7, 43.6, 41.2, 39.9, 38.6, 38.4, 37.8, 34.9, 28.7,
28.2, 19.5, 18.8, 18.5, 14.1, 13.6; HRMS (ESI, m/z) calcd for
C22H34O4Na [M+Na]+ 385.2355. Found: 385.2355.
4.3.12.1. Ethyl ent-15
(12a). Yield 91%; IR (KBr): 2954, 2851, 1745, 1722, 1460, 1381,
1236, 1180, 1152, 1124, 1097, 1032, 959, 604 cmÀ1 1H NMR
a-acetoxymethyl-16-oxobeyeran-19-oate
;
(400 MHz, CDCl3): d 4.37 (dd, J = 11.3, 5.9 Hz, 1H), 4.22 (dd,
J = 11.3, 3.7 Hz, 1H), 4.10 (m, 2H), 2.62 (m, 1H), 2.17 (d,
J = 13.4 Hz, 1H), 2.03 (s, 3H), 1.86–1.71 (m, 8H), 1.51–1.26 (m,
4H), 1.25 (t, J = 7.1 Hz, 3H), 1.24–1.20 (m, 2H), 1.19 (s, 3H), 1.14–
0.98 (m, 2H), 0.97 (s, 3H), 0.95–0.74 (m, 2H), 0.70 (s, 3H); 13C
NMR (100 MHz, CDCl3): d 221.8, 177.1, 170.5, 62.1, 59.9, 56.9,
56.8, 52.8, 50.9, 48.0, 43.5, 40.6, 39.6, 38.2, 37.8, 37.1, 35.4, 28.8,
21.4, 20.9, 19.7, 19.5, 18.8, 14.1, 13.3; HRMS (ESI, m/z) calcd for
C25H38O5Na [M+Na]+ 441.2617. Found: 441.2612.
4.3.15. General procedure for synthesis of compounds 15a and
15b
A mixture of compound 1or 2 (1 mmol) and phenylhydrazine
(1.1 mmol) in acetic acid (20 mL), saturated with gaseous HCl at
20 °C, was quickly warmed to boiling. When all solids were dis-
solved refluxing was continued for 5 min. Then reaction mixture
was evaporated and distributed between water and CH2Cl2. Organ-
ic phase was washed with saturated NaCl aqueous solution, dried
with MgSO4 and evaporated. The residue was purified by column
chromatography on silica to give product 15.
4.3.12.2. Ethyl ent-15
oate (12b). Yield 71%; mp 103–105 °C; IR (KBr): 2928, 2854,
1744, 1722, 1462, 1376, 1296 cmÀ1 1H NMR (400 MHz, CDCl3): d
a-benzoyloxymethyl-16-oxobeyeran-19-
;
8.00 (d, J = 8.0 Hz, 2H), 7.57 (t, J = 7.1 Hz, 1H), 7.45 (t, J = 7.7 Hz,
2H), 4.64 (dd, J = 11.4, 5.0 Hz, 1H), 4.53 (dd, J = 11.4, 3.2 Hz, 1H),
4.09 (m, 2H), 2.73 (m, 1H), 2.22 (d, J = 13.6 Hz, 1H) 2.00–1.70 (m,
8H), 1.46–1.28 (m, 10H), 1.25 (t, J = 7.2 Hz, 3H), 1.05 (s, 3H),
1.02–0.93 (m, 2H), 0.76 (s, 3H); 13C NMR (100 MHz, CDCl3): d
220.6, 176.4, 165.7, 133.7, 129.5, 129.2, 129.2, 129.0, 129.0, 62.7,
59.7, 56.2, 56.1, 52.2, 50.9, 47.7, 43.2, 40.3, 39.1, 37.9, 37.4, 36.6,
35.1, 28.6, 21.4, 20.0, 19.3, 18.7, 14.1, 13.1; HRMS (ESI, m/z) calcd
for C30H40O5Na [M+Na]+ 503.2773. Found: 503.2775.
4.3.15.1. Indole derivative of isosteviol (15a). Yield 80%; mp
154–156 °C; IR (KB): 3366, 3060, 2936, 2846, 1694, 1610, 1448,
1356, 1260, 1225, 1180, 1150, 1021, 970, 740 cmÀ1 1H NMR
;
(400 MHz, CDCl3): d 7.79 (s, 1H), 7.66 (d, J = 7.8 Hz, 1H), 7.27 (s,
1H), 7.05 (m, 1H), 7.01 (m, 1H), 2.58 (m, 1H), 2.21–1.77 (m, 6H),
1.66–1.35 (m, 5H), 1.33 (s, 3H), 1.28 (s, 3H), 1.26 (t, J = 7.2 Hz,
3H), 1.19–0.56 (m, 3H), 0.49 (s, 3H); 13C NMR (100 MHz, CDCl3):
d 179.4, 148.9, 138.7, 124.3, 119.3, 118.8, 118.3, 118.2, 111.5,
66.9, 56.4, 53.4, 44.9, 43.0, 39.6, 37.8, 37.1, 34.8, 30.5, 29.1, 28.8,
23.1, 22.2, 20.9, 18.9, 13.1; HRMS (ESI, m/z) calcd for C26H33NO2Na
[M+Na]+ 414.2409. Found: 414.2426.
4.3.13. Ethyl ent-15-methylene-16-oxobeyeran-19-oate (13)
A mixture of the compound 12 (1 mmol) and DBU (0.228 g,
1.5 mmol) in pyridine (15 mL) was stirred at 80 °C for 6 h, then
the reaction mixture was poured into water and acidified to pH 6
with 1 M hydrochloric acid. The aqueous layer was extracted with
CHCl3 and the filtrate was concentrated. The residue was purified
by column chromatography on silica (petroleum ether/ethyl ace-
tate = 10:1, v/v) to give product 13. Yield 85%; mp 109–111 °C; IR
(KBr): 3478, 2954, 2927, 2853, 1727, 1631, 1456, 1378, 1233,
4.3.15.2. Indole derivative of isosteviol ethyl ester (15b). Yield
91%; mp 165–167 °C; IR (KBr): 3381, 3048, 2944, 2839, 1720,
1608, 1446, 1374, 1230, 1149, 1091, 1022, 744, 513 cmÀ1 1H
;
NMR (400 MHz, CDCl3): d 7.81 (m, 1H), 7.67 (m, 1H), 7.31 (d,
J = 8.8 Hz, 1H), 7.06 (m, 2H), 4.17 (m, 2H), 2.43 (m, 1H), 2.19–
1.78 (m, 5H), 1.73–1.53 (m, 7H), 1.40 (t, J = 7.2 Hz, 3H), 1.28 (s,
3H), 1.23 (s, 3H), 1.19–0.71 (m, 5H), 0.47 (s, 3H); 13C NMR (100
MHz, CDCl3): d 177.8, 148.6, 138.8, 124.9, 121.0, 119.7, 119.6,
119.5, 111.6, 67.3, 60.1, 57.1, 54.0, 45.4, 44.0, 40.5, 40.4, 38.4,
37.5, 37.3, 35.1, 28.9, 23.5, 22.3, 21.4, 19.2, 14.1, 13.4; HRMS
(ESI, m/z) calcd for C28H37NO2Na [M+Na]+ 442.2722. Found:
442.2723.
1177, 1151, 1110, 1028 cmÀ1 1H NMR (400 MHz, CDCl3): d 5.90
;
(s, 1H), 5.39 (s, 1H), 4.03 (m, 2H), 2.07–2.02 (m, 2H), 1.93–1.87
(m, 1H), 1.72–1.46 (m, 7H), 1.37–1.17 (m, 6H), 1.16 (s, 3H), 1.14–
1.00 (m, 3H), 0.92 (s, 3H), 0.90–0.70 (m, 2H), 0.55 (s, 3H); 13C
NMR (100 MHz, CDCl3): d 210.8, 177.2, 154.5, 115.8, 60.0, 56.8,
56.6, 54.6, 54.2, 53.4, 48.4, 46.7, 43.7, 41.5, 39.4, 38.0, 37.3, 28.9,
21.7, 20.3, 19.6, 14.1, 12.4; HRMS (ESI, m/z) calcd for C23H34O3Na
[M+Na]+ 381.2406. Found: 381.2407.
4.3.16. General procedure for synthesis of compounds 16a and
16b
4.3.14. General procedure for synthesis of compounds 14a and
14b
A mixture of compound 1or 2 (1 mmol) and m-CPBA (0.258 g,
1.5 mmol) in CH2Cl2 was stirred at 0 °C for 5 h, then the reaction
mixture was poured into water and neutralized with NaHCO3
aqueous solution. The aqueous layer was extracted with CH2Cl2
A mixture of compound 1or 2 (1 mmol) and hydroxylamine
hydrochloride (0.103 g, 1.5 mmol) in C2H5OH was stirred in pres-
ence of NaHCO3 at 60 °C for 2 h, then the reaction mixture was con-
centrated under vacuum, and extracted with CH2Cl2 and H2O. At
last the organic layer was washed with saturated NaCl aqueous

1472
Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
solution, dried with MgSO4 and concentrated under vacuum to
give white powder 16.
was concentrated under vacuum , and extracted with CH2Cl2 and
H2O. At last the organic layer was washed with saturated NaCl
aqueous solution, dried with MgSO4 and concentrated under vac-
uum. The residue was purified by column chromatography on
silica to give products 19 and 20.
4.3.16.1. ent-16-Hydroximinobeyeran-19-oic acid (16a)27
.
Yield 90%; mp 103–104 °C; IR (KBr): 3425, 3118, 2942, 2847,
1694, 1471, 1454, 1417, 1387, 1250, 1215, 1188, 1153, 946,
795 cmÀ1
;
1H NMR (400 MHz, CDCl3): d 2.17 (d, J = 9.5 Hz, 1H),
4.3.18.1. Lactam of isosteviol (19a)37
3389, 3213, 2927, 2849, 1713, 1632, 1466, 1396, 1261, 1226,
1171, 1153, 991, 971, 782 cmÀ1 1H NMR (400 MHz, CDCl3): d
6.44 (s, 1H), 3.00 (d, J = 18.4 Hz, 1H), 2.18 (d, J = 13.2 Hz, 1H),
2.03 (d, J = 18.8 Hz, 1H), 1.96–1.77 (m, 4H), 1.70–1.57 (m, 3H),
1.50–1.38 (m, 2H), 1.30–1.25 (m, 4H), 1.22 (s, 3H), 1.20 (s, 3H),
1.08–0.89 (m, 3H), 0.88 (s, 3H), 0.86–0.80 (m, 1H); 13C NMR (100
MHz, CDCl3): d 181.3, 176.6, 58.5, 57.9, 52.8, 50.2, 49.3, 45.2,
44.7, 41.1, 40.9, 40.6, 39.1, 39.0, 36.2, 29.3, 28.5, 20.9, 20.1, 19.9;
HRMS (ESI, m/z) calcd for C20H31NO3Na [M+Na]+ 356.2202. Found:
356.2199.
. Mp 98–100 °C; IR (KBr):
2.13 (d, J = 15.1 Hz, 1H), 2.05–1.68 (m, 4H), 1.65 (d, J = 1.4 Hz,
3H), 1.62–1.29 (m, 7H), 1.23 (s, 3H), 1.14–1.18 (m, 3H), 0.96 (s,
3H), 0.87 (s, 3H), 0.86–0.78 (m, 1H); HRMS (ESI, m/z) calcd for
C20H32NO3 [M+H]+ 334.2382. Found: 334.2387.
;
4.3.16.2. Ethyl ent-16-hydroximinobeyeran-19-oate (16b).
Yield 95%; mp 42–44 °C; IR (KBr): 3306, 2939, 2847, 1723, 1450,
1377, 1320, 1299, 1233, 1179, 1152, 1097, 1028, 929, 849, 792,
722, 575 cmÀ1 1H NMR (400 MHz, CDCl3): d 4.08 (m, 2H), 2.97
;
(d, J = 18.6 Hz, 1H), 2.17 (d, J = 13.3 Hz, 1H), 2.00 (d, J = 18.6 Hz,
1H), 1.89–1.57 (m, 7H), 1.48–1.39 (m, 4H), 1.26 (t, J = 7.1 Hz, 3H),
1.23–1.20 (m, 2H), 1.18 (s, 3H), 1.10 (s, 3H), 1.09–0.84 (m, 4H),
0.77 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.4, 170.4, 59.9,
57.0, 56.2, 54.8, 43.7, 43.6, 40.8, 40.5, 39.8, 39.3, 38.0, 37.9, 36.7,
28.8, 22.0, 21.6, 20.3, 18.8, 14.1, 13.2; HRMS (ESI, m/z) calcd for
C22H35NO3Na [M+Na]+ 384.2515. Found: 384.2512.
4.3.18.2. Lactam of isosteviol ethyl ester (19b). Mp 65–67 °C; IR
(KBr): 3430, 3199, 2946, 2847, 1722, 1661, 1471, 1328, 1233, 1149,
1019 cmÀ1; 1H NMR (400 MHz, CDCl3): d 6.24 (s, 1H), 4.09 (m, 2H),
2.90 (d, J = 18.4 Hz, 1H), 2.16 (d, J = 13.2 Hz, 1H), 1.94 (d, J = 18.3
Hz, 1H), 1.90–1.75 (m, 4H), 1.63–1.46 (m, 3H), 1.41–1.27 (m, 3H),
1.26 (t, J = 7.2 Hz, 3H), 1.18 (s, 3H), 1.17 (s, 3H), 1.09–0.84 (m,
4H), 0.80 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.1, 173.9,
60.0, 57.4, 56.7, 51.7, 49.3, 44.2, 43.6, 40.2, 39.6, 39.5, 37.9, 37.8,
35.1, 28.8, 28.6, 19.7, 18.8, 18.7, 14.1, 13.7; HRMS (ESI, m/z) calcd
for C22H33NO2Na [M+Na]+ 366.2409. Found: 366.2403.
4.3.17. General procedure for synthesis of compounds 17 and 18
Compound 16a or 16b (1 mmol) was dissolved in C2H5OH or
THF (30 mL) followed by addition of Ni (0.5 mmol) in catalytic
hydrogenation flask. The reaction proceeded 4 h under 3 atm
hydrogen at 40 °C. Then the reaction mixture was filtered, and
the filtrate was concentrated. The residue was purified by column
chromatography on silica to give products 17 and 18.
4.3.18.3. Mixture of compounds 20a and 20b. A mixture of 20a
and 20b isomers (3:1 determined by 1H NMR); IR (KBr): 3002,
2950, 2922, 2858, 2239, 1717, 1454, 1377, 1227, 1147 cmÀ1 1H
;
4.3.17.1. ent-16-Aminobeyeran-19-oic acid (17a). Yield 82%;
mp 367–368 °C; IR (KBr): 3428, 2940, 2846, 2678, 1689, 1616,
NMR (400 MHz, CDCl3): d 5.36 (s, 1H), 4.09 (m, 2H), 2.76–1.74
(m, 10H), 1.66 (s, 3H), 1.60–1.34 (m, 5H), 1.26 (t, J = 7.2 Hz, 3H),
1.19 (s, 3H), 1.17–0.71 (m, 3H), 0.68 (s, 3H); 13C NMR (100 MHz,
CDCl3): for 20a: d 177.1, 131.2, 119.8, 119.0, 60.1, 57.1, 51.7,
48.4, 45.7, 43.6, 39.7, 39.0, 37.7, 37.3, 35.2, 28.7, 23.3, 22.1, 20.0,
19.7, 18.9, 14.1, 13.5; HRMS (ESI, m/z) calcd for C22H35NO3Na
[M+Na]+ 384.2515. Found: 384.2514.
1546, 1464, 1398 cmÀ1 1H NMR (400 MHz, DMSO): d 3.48 (m,
;
1H), 2.17 (d, J = 13.3 Hz, 1H), 1.99–1.57 (m, 8H), 1.48–1.39 (m,
5H), 1.27 (s, 3H), 1.23–1.10 (m, 4H), 1.08 (s, 3H), 1.07–0.88 (m,
3H), 0.87 (s, 3H); HRMS (ESI, m/z) calcd for C20H34NO2 [M+H]+
320.2590. Found: 320.2576.
4.3.17.2. Ethyl ent-16-aminobeyeran-19-oate (17b). Yield 88%;
mp 89–91 °C; IR (KBr): 3353, 2939, 2845, 1722, 1661, 1451,
4.4. X-ray crystallographic analysis
1374, 1231, 1177, 1151, 1048 cmÀ1
;
1H NMR (400 MHz, CDCl3): d
X-ray crystal data of compounds 4, 6, 10f, 12b and 14b were
4.08 (q, J = 7.2 Hz, 2H), 2.89 (dd, J = 11.2, 6.0 Hz, 1H), 2.15 (d,
J = 13.2 Hz, 1H), 1.85–1.51 (m, 9H), 1.40–1.31 (m, 6H), 1.26 (t,
J = 7.2 Hz, 3H), 1.16 (s, 3H), 1.04–0.85 (m, 4H), 0.84 (s, 3H), 0.73
(s, 3H); 13C NMR (100 MHz, CDCl3): d 177.5, 61.0, 59.8, 57.1, 56.5,
55.6, 43.6, 42.9, 41.8, 41.4, 40.0, 39.9, 37.9, 34.2, 33.3, 28.9, 24.8,
21.7, 20.5, 18.8, 14.0, 13.3; HRMS (ESI, m/z) calcd for C22H38NO2
[M+H]+ 348.2903. Found: 348.2904.
collected by a Rigaku AFC5R diffractometer with graphite-mono-
chromated Mo-Ka radiation (k = 0.71073 Å). The structure was
solved by the direct method and refined with a full-matrix least
squares method.
4.4.1. Crystal data for compound 4
C22H36O3, M = 348.51, orthorhombic, space group P212121,
a = 7.5578(4), b = 15.1297(8), c = 17.2037(10), V = 1967.20(19) Å3,
4.3.17.3. Ethyl ent-16-aminoethylbeyeran-19-oate (18). Yield
86%; mp 61–63 °C; IR (KBr): 3433, 2936, 2844, 1723, 1454, 1377,
Z = 4,
tal dimensions: 0.41 Â 0.23 Â 0.12 mm, A total of 15,181 reflec-
tions (3650 unique) were collected using the -2h scan
technique to a maximum 2h value of 51°, and 2948 reflections with
I > 2 (I) were used in the structure determination. Final R and Rw
l(Mo-Ka
) = 0.076 cmÀ1, F(000) = 768, Dc = 1.177 g/cm3, crys-
1231, 1177, 1147, 1027, 973 cmÀ1
;
1H NMR (400 MHz, CDCl3): d
x
4.08 (m, 2H), 2.73–2.61 (m, 3H), 2.15 (d, J = 13.2 Hz, 1H), 1.82–
1.77 (m, 2H), 1.68–1.50 (m, 8H), 1.36–1.32 (m, 4H), 1.25 (t,
J = 7.2 Hz, 3H), 1.16 (s, 3H), 1.09 (t, J = 7.2 Hz, 3H), 1.04–0.94 (m,
5H), 0.91 (s, 3H), 0.73 (s, 3H); 13C NMR (100 MHz, CDCl3): d 177.5,
67.0, 59.8, 57.2, 57.0, 55.9, 43.6, 43.6, 42.1, 41.8, 41.4, 40.0, 39.9,
38.0, 38.0, 34.0, 28.9, 25.7, 21.7, 20.7, 18.9, 15.6, 14.1, 13.4; HRMS
(ESI, m/z) calcd for C24H42NO2 [M+H]+ 376.3216. Found: 376.3215.
r
values were 0.048 and 0.112, respectively. The maximum and min-
imum peaks in the difference map were 0.368 and À0.244 e ÅÀ3
,
respectively. The data have been deposited with the Cambridge
Crystallographic Data Centre as supplementary publication num-
ber CCDC 654209.
4.3.18. General procedure for synthesis of compounds 19 and
20
4.4.2. Crystal data for compound 6
C22H38O5, M = 382.52, monoclinic, space group P21, a = 2.151(2),
A mixture of the compound 16 (1 mmol) and BF3ÁOEt2 (30 mL)
b = 7.3549(15), c = 12.764(3), V = 1039.7(4) Å3, Z = 2,
l
(Mo-Ka) =
in toluene was stirred at 100 °C for 12 h, then the reaction mixture
0.084 cmÀ1, F(000) = 420, Dc = 1.222 g/cm3, crystal dimensions:

Y. Wu et al. / Bioorg. Med. Chem. 17 (2009) 1464–1473
1473
0.22 Â 0.20 Â 0.20 mm, A total of 2705 reflections (2641 unique)
were collected using the -2h scan technique to a maximum 2h
value of 49°, and 1883 reflections with I > 2 (I) were used in the
structure determination. Final R and Rw values were 0.0603 and
0.1475, respectively. The maximum and minimum peaks in the dif-
ference map were 0.720 and À0.407 e ÅÀ3, respectively. The data
have been deposited with the Cambridge Crystallographic Data
Centre as supplementary publication number CCDC 257291.
References and notes
x
1. Kameda, Y.; Asano, N.; Yoshikawa, M.; Takeucki, M.; Yamaguchi, T.; Matsui, K.;
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4.4.3. Crystal data for compound 10f
C25H42O4, M = 406.59, orthorhombic, space group P212121,
a = 7.8760(16), b = 20.434(4), c = 29.280(6), V = 4712.3(16) Å3, Z =
8,
dimensions: 0.20 Â 0.18 Â 0.17 mm, A total of 12,546 reflections
(7189 unique) were collected using the -2h scan technique to a
maximum 2h value of 50°, and 5249 reflections with I > 2 (I) were
l(Mo-Ka
) = 0.075 cmÀ1, F(000) = 1792, Dc = 1.146 g/cm3, crystal
x
r
used in the structure determination. Final R and Rw values were
0.0709 and 0.1778, respectively. The maximum and minimum
peaks in the difference map were 0.767 and À0.733 e ÅÀ3, respec-
tively. The data have been deposited with the Cambridge Crystallo-
graphic Data Centre as supplementary publication number CCDC
705071.
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4.4.4. Crystal data for compound 12b
C30H40O5, M = 480.62, orthorhombic, space group P212121,
a = 7.6199(15), b = 12.413(3), c = 27.789(6), V = 2628.5(9) Å3, Z = 4,
l
(Mo-K
dimensions: 0.30 Â 0.20 Â 0.20 mm, a total of 9122 reflections
(3197 unique) were collected using the -2h scan technique to a
maximum 2h value of 55°, and 2328 reflections with I > 2 (I) were
a
) = 0.081 cmÀ1, F(000) = 1040, Dc = 1.215 g/cm3, crystal
x
r
used in the structure determination. Final R and Rw values were
0.0588 and 0.1251, respectively. The maximum and minimum peaks
in the difference map were 0.279 and À0.217 e ÅÀ3, respectively.
The data have been deposited with the Cambridge Crystallographic
Data Centre as supplementary publication number CCDC 257292.
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4.4.5. Crystal data for compound 14b
C
22H34O4, M = 362.49, orthorhombic, space group P212121,
a = 6.9969(14), b = 12.671(3), c = 23.212(5), V = 2057.9(7) Å3, Z = 4,
(Mo-K
) = 0.079 cmÀ1 F(000) = 792, Dc = 1.170 g/cm3, crystal
dimensions: 0.20 Â 0.17 Â 0.17 mm, a total of 6421 reflections
(2165 unique) were collected using the -2h scan technique to a
maximum 2h value of 51°, and 1884 reflections with I > 2 (I) were
l
a
,
x
r
used in the structure determination. Final R and Rw values were
0.0576 and 0.1475, respectively. The maximum and minimum
peaks in the difference map were 0.241 and À0.203 e ÅÀ3, respec-
tively. The data have been deposited with the Cambridge Crystallo-
graphic Data Centre as supplementary publication number CCDC
705070.
33. Donoruma, L. G.; Heldt, V. Z.. In Organic Reactions; Wiley: New York, 1960; Vol.
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E.; Alfonsov, V. A.; Dmitry, R. Z.; Beskrovny, D. V.; Litvinov, I. A. Mendeleev
Commun. 2005, 15, 27.
Acknowledgment
The authors are grateful for the financial support to the National
Natural Science Foundation of China (Project No. 20772113).