Synthesis and Evaluation of a Series of Oleanolic Acid Saponins as α-Glucosidase and α-Amylase Inhibitors

Article abstract of DOI:10.1002/ardp.201500179

Sixteen naturally occurring oleanolic acid saponins and their derivatives were synthesized in an efficient and practical strategy, and their inhibitory activities against α-glucosidase and α-amylase were evaluated in vitro. Among all the compounds, 28-O-m

Full text of DOI:10.1002/ardp.201500179

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A P
Arch. Pharm. Chem. Life Sci. 2015, 348, 615–628
Archiv der Pharmazie
Full Paper
Synthesis and Evaluation of a Series of Oleanolic Acid Saponins
as a-Glucosidase and a-Amylase Inhibitors
Tiantian Guo1, Shaoping Wu2, Sen Guo1, Lu Bai1, Qingchao Liu1,3, and Naisheng Bai1*
1
Department of Pharmaceutical Engineering, Northwest University, Xi’an, Shaanxi, P. R. China
Sorbonne Universites, UPMC University, Paris, France
Department of Applied Biology and Chemical Technology, The Hong Kong Polytechnic University,
Kowloon, Hong Kong, P. R. China
2
ꢀ
3
Sixteen naturally occurring oleanolic acid saponins and their derivatives were synthesized in an
efficient and practical strategy, and their inhibitory activities against a-glucosidase and a-amylase
were evaluated in vitro. Among all the compounds, 28-O-monoglucoside 8 exhibited remarkably
potent inhibitory activity against a-glucosidase with an IC50 value of 87.3 mM, which was fivefold
stronger than that of the antidiabetic acarbose. Based on the preliminary structure–activity
relationships, for 28-O-monoglucosides, the presence of a terminal a-L-rhamnopyranosyl residue
enhanced the a-glucosidase and a-amylase inhibitory activities. Furthermore, for 3,28-O-bidesmosides,
sugar-substituted moieties attached to the C-3 and C-28 positions of the oleanolic acid scaffold are
helpful to increase the inhibitory activities against a-amylase and a-glucosidase.
Keywords: a-Amylase / a-Glucosidase / Glycosylation / Oleanolic acid saponin / Synthesis
Received: May 27, 2015; Revised: July 4, 2015; Accepted: July 7, 2015
DOI 10.1002/ardp.201500179
Additional supporting information may be found in the online version of this article at the publisher’s web-site.
:
level after meals, which has emerged as an important method
to control blood sugar levels [3–6].
Intestinal a-glucosidase, a membrane-bound enzyme in-
Introduction
Diabetes mellitus, which is a metabolic disorder characterized
by high plasma glucose levels, is the main threat to human
health all over the world [1]. Hyperglycemia, which is the
result of decreased insulin sensitivity or reduced insulin
secretion from pancreatic b-cells, can further inhibit insulin
secretion from pancreas and diminish insulin-mediated
glucose uptake in peripheral tissues [2]. Controlling plasma
glucose levels is essential for delaying or preventing type 2
diabetes. The inhibition of a-glucosidase and a-amylase can
extremely reduce the postprandial increase of blood glucose
volved in intestinal absorption, which is located in the brush-
border surface membrane of intestinal cells, hydrolyses the
cleavage of glucose from disaccharides and oligosaccharides
[7–10]. Pancreatic a-amylase, one of the major secretory
products of the pancreas and salivary glands, catalyzes the
hydrolysis of a-(1,4)-glucosidic linkages and produces maltose
and glucose from starch [11–14]. Consequently, inhibitors of
these hydrolytic enzymes can delay the absorption of glucose
into the bloodstream and prevent sudden postprandial
hyperglycemia after a mixed carbohydrate diet.
Considerable efforts have been made in the development
of small-molecule a-glucosidase and a-amylase inhibitors for
the treatment of type
2 diabetes [15–22]. In fact, a-
glucosidase and a-amylase inhibitors, such as voglibose,
Correspondence: Dr. Qingchao Liu, Department of Pharmaceutical
Engineering, Northwest University, Xi’an 710069, Shaanxi,
P. R. China.
E-mail: liuqc21@nwu.edu.cn
Fax: þ86-029-88305682
ꢀAdditional correspondence: Prof. Naisheng Bai,
E-mail: nsbai@nwu.edu.cn
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miglitol, and acarbose effectively compensate for defective
early-phase insulin release by inhibiting postprandial absorp-
tion of monosaccharides, and have been approached for
clinical use to the management of type 2 diabetes [23–27].
However, these drugs are often associated with undesirable
side-effects, such as adverse gastrointestinal symptoms (like
abdominal flatulence, diarrhea, and discomfit) and liver
toxicity, which depress patient compliance and treatment
effectiveness [28, 29]. Therefore, there is a need for potent
and effective a-glucosidase and a-amylase inhibitors with no
adverse or unwanted secondary effects.
Triterpenoid saponins are glycosylated plant secondary
metabolites, which have been reported to possess many
important biological and pharmacological activities [30–33].
In our research program for potent and effective antidiabetic
agents, carbohydrate-based modifications on natural products
(especiallytriterpenoid saponins) werecarriedout. In a seriesof
our studies on in vitro PTP1B and a-glucosidase-inhibiting
principles from synthetic natural products and derivatives, we
previously reported the efficient artificial synthesis of sugar-
substituted oleanolic acid derivatives, hederagenin glycosides,
natural benzophenone O-glycosides, and their derivatives. The
structure–activity relationships (SARs) of these saponins
showed that the sugar moiety strongly influenced PTP1B and
a-glucosidase inhibitory activities, and also improved their
inhibition against murine in vivo glucoseabsorption [34–37]. As
part of an ongoing investigation, here we report the synthesis,
a-glucosidase and a-amylase inhibitory activities, and SARs of
naturally occurring oleanane-type triterpenoid saponins (1, 5–
13, 15, and 16) [38–43] and non-natural oleanolic acid saponins
(2–4 and 14) (Fig. 1).
donors 7a–9a (Scheme 2) involved in the synthesis were
readily obtained by one step bromination reaction. So, the full
benzoylated glycosides were brominated with hydrobromic
acid in glacial acetic acid giving glycosyl bromide donors 7a–
9a [44]. Treatment of oleanolic acid with glycosyl bromide
donor (7a, 8a, or 9a) under the modified literature conditions
(K2CO3, TBAB, CH2Cl2/H2O, reflux) [45, 46] afforded the
desired C28-O-glycosyl esters 6b–8b in good yields. Finally,
removal of the benzoyl groups with NaOMe in MeOH/CH2Cl2
afforded the target compounds 6–8 in satisfactory yields.
Oleanolic acid C3-O-glycosides 9–16 were synthesized as
shown in Scheme 3. The glycosyl trichloroacetimidate donors
3a–6a and 10a–13a (Schemes 1 and 3) were coupled with
oleanolic ester 17 in the presence of TMSOTf (0.3 equiv.) at
ꢁ78°C for 30 min providing the key intermediates, which were
then transformed into the desired glycosides 9b–16b by
warming to room temperature for 30 min. After removal of all
the benzoyl groups with NaOMe in MeOH/CH2Cl2, the
glycosides 9b–16b were transformed into the target com-
pounds 9–16 in satisfactory yields.
Biological evaluations
Theinhibitoryeffectofsaponins1–16againsta-glucosidasewas
screened using p-nitrophenyl-a-glucopyranoside (pNPG) as a
substrate, and the a-amylase inhibitory effect was also deter-
mined using p-nitrophenyl-a-maltopentoglycoside (pNPM).
The commercial a-glucosidase inhibitor, acarbose, was chosen
aspositivecontrolandreferencepointintheassays.Asshownin
Table 1, compounds 6–8 with 28-O-monoglucosides showed
more potent a-glucosidase and a-amylase inhibitory activities
thanthatofacarbose.Notably,compound8(IC50,87.3 mM)with
a terminal a-L-rhamnopyranosyl residue exhibited the stronger
a-glucosidase inhibitory activity, compared with compounds 6
(IC50, 251.7 mM) and 7 (IC50, 200.7 mM). This result indicates that
the spatial configuration of a terminal a-L-rhamnopyranosyl
residue is favorable to the a-glucosidase inhibitory activity. In
previous reports, the monodesmosidic saponin 7 exhibited
cytotoxic activity against human tumor HL-60, HepG-2, and
SGC-7901 cell lines [41], and compound 8 upregulated PPARg
transcriptional activity in a dose-dependent manner in HepG2
cells [42]. However, their a-amylase and a-glucosidase inhibito-
ry effects have not been reported. Compounds 9–16 with 3-O-
monoglucosides were found to have no inhibitory activities
Results and discussion
Chemistry
The synthetic route of bidesmosidic oleanolic acid saponins 1–
5 is shown in Scheme 1. According to our previously reported
one-pot protocols [36], saponins 1–5 were synthesized with
two glycose 1-(trichloroacetimidate)s as donors. The glycosyl
trichloroacetimidate donors 1a–6a (Scheme 1) involved in the
synthesis were readily prepared according to the previously
reported procedures [34–36]. Condensation of oleanolic ester
17 with glycosyl trichloroacetimidate donor (1a, 2a, or 3a) in
the presence of a catalytic amount of trimethylsilyl trifluor-
omethanesulfonate (TMSOTf, 0.3 equiv.) at ꢁ78°C for 30 min
obtained the desired glycoside, which was then transformed
into the key intermediate by warming to room temperature
for 30 min. After addition of a CH2Cl2 solution of the glycosyl
trichloroacetimidates donor (4a, 5a, or 6a) to the above
mixture at 0°C, the desired glycosides (1b–5b) were obtained
in satisfactory yields. Subsequent deprotection of the acetyl
and benzoyl esters with NaOMe in MeOH/CH2Cl2 was
achieved to afford the target compounds 1–5 in good yields.
Oleanolic acid C28-O-glycosides 6–8 were prepared accord-
ing to the procedure in Scheme 2. The glycosyl bromide
against a-glucosidase, and interestingly, compounds 15 (IC50
,
356.3 mM) and 16 (IC50, 410.6 mM) exhibited higher inhibitory
activity against a-amylase than compounds 9–14, indicating
that the disaccharide linkage attached to 3-OH of the aglycone
resulted in an increase in a-amylase inhibitory activity.
Compared with acarbose, compounds 1–5 with 3,28-O-bides-
mosides showed similar a-glucosidase anda-amylase inhibitory
activities.
Actually, Yoshikawa and co-workers reported that olea-
nolic acid 28-O-b-D-glucopyranoside 6 lacked the inhibitory
effect on the increase in serum glucose in oral glucose-loaded
rats. However, oleanolic acid 3-O-b-D-glucopyranoside
9
exhibited significant hypoglycemic activity [40]. Compared
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Arch. Pharm. Chem. Life Sci. 2015, 348, 615–628
Oleanolic Acid Saponins as a-Glucosidase Inhibitors
Archiv der Pharmazie
Figure 1. Chemical structures of oleanolic acid saponins (1–16).
with these reported results, our experimental results indicat-
ed that compounds 6 and 9 may adopt different action modes
on hypoglycemic activity [40]. Based on our results, the
preliminary SARs demonstrated that 28-O-monoglucosides
(6–8) exhibited potent a-glucosidase and a-amylase inhibitory
activities, which is consistent with the results reported by Luo
and co-workers [30]. Furthermore, 3,28-O-bidesmosides (1–5)
and positive control (acrbose) showed similar a-glucosidase
and a-amylase inhibitory activities, and 3-O-monoglucosides
(9–16) showed no inhibitory activities against a-glucosidase.
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Scheme 1. Synthetic route for compounds 1–5. Reagents and conditions: (a) TMSOTf (0.3 equiv.), CH2Cl2, ꢁ78°C, 30 min; room
temperature, 30 min; (b) TMSOTf (0.1 equiv.), CH2Cl2, 0°C, 30 min, 61–68% for 1b–5b; (c) NaOMe, MeOH/CH2Cl2, 89–93% for 1–5.
improve pharmacological profile and clarify action modes for
this series of sugar-subsitituted oleanolic acid are currently
Conclusions
In conclusion, we have synthesized a series of natural and non-
natural oleanolic acid saponins in an efficient and practical
way, and their a-glucosidase and a-amylase inhibitory
activities were evaluated in vitro. On the basis of preliminary
structure–activity relationships, we could summarize that: (i)
for 28-O-monoglucosides, the presence of a terminal a-L-
rhamnopyranosyl residue enhances inhibitory activity; (ii) for
3-O-monoglucosides, the disaccharide linkage attached to 3-
OH of the aglycone increases a-amylase inhibitory activity; (iii)
for 3,28-O-bidesmosides, sugar-substituted moieties attached
to the C-3 and C-28 positions of OA scaffold are favorable to
enhance the inhibitory activity against a-amylase and a-
glucosidase inhibitory activities. Further investigations to
under way in our research group and will be reported in due
time.
Experimental
Chemistry
Commercial reagents were used without further purification
unless specialized. Solvents were dried and redistilled prior to
use in the usual way. Thin-layer chromatography (TLC) was
performed on precoated E. Merck Silica Gel 60 F254 plates.
Flash column chromatography was performed on silica gel
(200–300 mesh). Optical rotations were determined with a
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Arch. Pharm. Chem. Life Sci. 2015, 348, 615–628
Oleanolic Acid Saponins as a-Glucosidase Inhibitors
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Scheme 2. Synthetic route for compounds 6–8. Reagents and conditions: (a) K2CO3, TBAB, CH2Cl2/H2O, reflux, 89% for 6b, 85% for
7b, 87% for 8b; (b) NaOMe, MeOH/CH2Cl2, 95% for 6, 89% for 7, 87% for 8.
Perkin–Elmer Model 241 MC polarimeter. 1H NMR and 13C
NMR spectra were taken on a Jeol JNM-ECP 600 spectrometer
with tetramethylsilane as the internal standard, and chemical
shifts are recorded in d values. Mass spectra were recorded on
a Q-TOF Global mass spectrometer.
purified by a silica gel column chromatography (petroleum
ether/EtOAc) to afford the products 1b–5b.
3b-O-(2,3,4,6-Tetra-O-benzoyl-b-D-glucopyranosyl)-(1!4)-
2,3,6-tri-O-benzoyl-b-D-glucopyranosyl)oleanolic acid,
2,3,4,6-tetra-O-acetyl-b-D-glucopyranosyl ester (1b)
23
Typical procedure for the synthesis of compounds 1b–5b
A mixture of compound 17 (100 mg, 0.11 mmol), trichloroacet-
imidate 1a, 2a, or 3a (0.14 mmol, 1.2 equiv.), and powdered
Yield: 65%; [a]D þ 31.3 (c 1.34, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.18–8.03 (m, 35 H, Ph-H), 5.77 (t, J ¼ 10.0 Hz, 1 H,
H-300), 5.61 (t, J ¼ 9.7 Hz, 1H, H-30), 5.54 (d, J ¼ 8.2 Hz, 1 H, H-
1000), 5.43 (t, J ¼ 9.6 Hz, 1 H, H-3000), 5.29 (t, J ¼ 3.2 Hz, 1 H, H-12),
5.23 (t, J ¼ 9.7 Hz, 1 H, H-400), 5.18 (t, J ¼ 9.6 Hz, 1 H, H-40), 5.13
(d, J ¼ 9.6 Hz, 1H, H-4000), 4.94 (d, J ¼ 7.6 Hz, 1 H, H-100), 4.68 (d,
J ¼ 7.6 Hz, 1H, H-10), 4.57 (dd, J ¼ 9.6, 7.6 Hz, 1 H, H-2000), 4.48
(dd, J ¼ 10.0, 8.3 Hz, 1 H, H-2000), 4.24–4.32 (m, 2 H, H-20, H-6000-
1), 4.09–4.11 (m, 2 H, H-50, H-5000), 3.99–4.08 (m, 2 H, H-600-1, H-
6000-2), 3.70-3.81 (m, 3 H, H-60-1, H-60-2, H-600-2), 2.95 (dd,
J ¼ 11.2, 4.4 Hz, 1 H, H-3), 2.78 (dd, J ¼ 13.7, 3.7 Hz, 1 H, H-18),
2.04, 2.01, 1.99, 1.97 (s each, 3 H each, Ac ꢂ 4), 1.06, 0.90, 0.90,
0.76, 0.64, 0.60, 0.56 (s each, 3 H each, CH3 ꢂ 7); 13C NMR
̊
4 A molecular sieves in dry CH2Cl2 (5 mL) were stirred for
30 min at room temperature and then cooled to 0°C. TMSOTf
(15 mL, 0.09 mmol, 0.3 equiv.) was added slowly. After being
stirred at ꢁ78°C for 30 min, the reaction mixture was warmed
up to room temperature for 30 min, and then cooled to 0°C. A
solution of 4a, 5a, or 6a (0.17 mmol, 1.5 equiv.) in dry CH2Cl2
(5 mL) was injected slowly. The reaction mixture was stirred at
0°C for 30 min, and then warmed up to room temperature for
another 30 min. The reaction was quenched by addition of
Et3N and then filtered. The filtrate was concentrated and
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Scheme 3. Synthetic route for compounds 9–16. Reagents and conditions: (a) TMSOTf (0.3 equiv.), CH2Cl2, ꢁ78°C, 30 min; room
temperature, 30 min, 81–89% for 9b–16b; (b) NaOMe, MeOH/CH2Cl2, 87–93% for 9–16.
(CDCl3, 100 MHz): d 175.6 (C-28), 170.6, 170.1, 169.4, 168.9
(Ac), 165.7, 165.1, 156.8, 142.8 (C-13), 133.3, 133.2, 133.0,
130.0, 129.9, 129.8, 129.7, 129.4, 128.8, 128.6, 128.5, 128.3,
128.2, 123.0 (C-12), 103.0 (C-1000), 100.9 (C-10), 91.5 (C-100), 90.6
(C-3), 72.8, 72.7, 72.4, 72.1, 71.9, 71.7, 71.5, 69.8, 69.3, 69.2,
69.1, 67.9, 62.6, 61.4, 60.4, 55.3, 47.4, 46.7, 45.7, 41.6, 41.0,
39.2, 38.6, 38.1, 36.5, 33.7, 33.0, 32.7, 31.7, 30.6, 27.5, 25.5,
23.4, 23.3, 20.6, 16.8, 16.1, 15.1, 14.2; HRMALDIMS: m/z calcd
for C105H114O29Na [MþNaþ] 1861.7338; found: 1861.7367.
H-1000), 5.53 (t, J ¼ 9.6 Hz, 1 H, H-3000), 5.48 (t, J ¼ 3.6 Hz, 1 H, H-
400), 5.38 (t, J ¼ 9.6 Hz, 1 H, H-40), 5.30 (t, J ¼ 3.6 Hz, 1 H, H-12),
5.23 (d, J ¼ 9.6 Hz, 1H, H-4000), 5.15 (d, J ¼ 7.9 Hz, 1 H, H-100), 4.89
(d, J ¼ 7.7 Hz, 1H, H-10), 4.57 (dd, J ¼ 9.6, 7.9 Hz, 1 H, H-200), 4.51
(dd, J ¼ 10.0, 8.0 Hz, 1 H, H-2000), 4.40 (dd, J ¼ 9.6, 7.7 Hz, 1 H, H-
20), 4.37 (dd, J ¼ 12.1, 3.5 Hz, 1 H, H-6000-1), 4.26–4.32 (m, 2 H, H-
5000, H-6000-2), 4.16 (m, 1 H, H-50), 4.07 (dd, J ¼ 12.3, 3.1 Hz, 1 H, H-
600-1), 3.85 (dd, J ¼ 12.0, 4.3 Hz, 1 H, H-60-1), 3.79 (dd, J ¼ 12.0,
5.7 Hz, 1 H, H-60-2), 4.37 (dd, J ¼ 12.3, 4.9 Hz, 1 H, H-600-2), 3.01
(dd, J ¼ 12.3, 3.7 Hz, 1 H, H-3), 2.81 (dd, J ¼ 13.7, 4.4 Hz, 1 H, H-
18), 2.05, 2.00, 1.99, 1.98 (s each, 3 H each, Ac ꢂ 4), 1.07, 0.93,
0.91, 0.75, 0.66, 0.60, 0.57 (s each, 3 H each, CH3 ꢂ 7); 13C NMR
(CDCl3, 100 MHz): d 175.7 (C-28), 170.9, 170.0, 169.7, 169.3
(Ac), 165.4, 165.2, 165.1, 142.8 (C-13), 133.5, 133.4, 133.2,
133.0, 130.0, 129.7, 129.6, 129.4, 129.3, 128.8, 128.7, 128.6,
128.5, 128.2, 122.8 (C-12), 103.1 (C-1000), 101.0 (C-10), 91.5 (C-
3b-O-(2,3,4,6-Tetra-O-benzoyl-b-D-galactopyranosyl)-
(1!4)-2,3,6-tri-O-benzoyl-b-D-glucopyranosyl)oleanolic
acid, 2,3,4,6-tetra-O-acetyl-b-D-glucopyranosyl ester (2b)
23
Yield: 67%; [a]D þ 20.5 (c 1.15, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.12–8.01 (m, 35 H, Ph-H), 5.81 (dd, J ¼ 10.0, 3.6 Hz,
1 H, H-300), 5.75 (t, J ¼ 9.6 Hz, 1H, H-30), 5.56 (d, J ¼ 8.0 Hz, 1 H,
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Oleanolic Acid Saponins as a-Glucosidase Inhibitors
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Table 1. Inhibitory activities of compounds 1–16 against a-glucosidase and a-amylase.
IC50 (mM)a)
IC50 (mM)a)
Compounds
a-Glucosidase
a-Amylase
Compounds
a-Glucosidase
a-Amylase
1
2
3
4
5
6
7
8
450.1 ꢃ 29.7
390.6 ꢃ 30.3
410.9 ꢃ 35.9
380.1 ꢃ 25.8
476.3 ꢃ 40.1
251.7 ꢃ 19.9
200.7 ꢃ 23.1
87.3 ꢃ 9.1
551.3 ꢃ 43.1
501.3 ꢃ 35.9
596.3 ꢃ 45.1
496.4 ꢃ 39.8
600.3 ꢃ 47.3
387.7 ꢃ 29.4
390.6 ꢃ 35.5
210.3 ꢃ 26.7
579.3 ꢃ 40.4
9
NAb)
NA
NA
NA
NA
NA
NA
NA
–
NA
NA
NA
NA
NA
NA
10
11
12
13
14
15
16
–
356.3 ꢃ 29.7
410.6 ꢃ 35.1
–
Acarbosec)
434.5 ꢃ 36.2
a) The IC50 values in mM were calculated from the dose response curve of six concentrations of each test compound in triplicate.
b) NA: not active.
c) Positive control.
100), 90.6 (C-3), 73.0, 72.8, 72.3, 72.0, 71.9, 71.4, 70.8, 69.9, 69.8,
67.9, 67.5, 62.7, 61.5, 61.1, 56.3, 47.4, 46.7, 45.7, 42.3, 41.6,
41.0, 39.3, 38.5, 38.1, 36.5, 33.6, 33.0, 32.9, 31.5, 30.6, 27.6,
25.7, 23.5, 23.3, 21.6, 16.8, 16.3, 15.1, 14.5; HRMALDIMS: m/z
38.6, 38.3, 38.0, 36.5, 33.6, 32.7, 31.6, 31.3, 30.4, 29.5, 27.8,
27.4, 25.6, 25.0, 23.4, 23.1, 22.3, 20.6, 17.9, 16.2, 16.0, 15.3;
HRMALDIMS: m/z calcd for
C
152H144O37Na [MþNaþ]
2583.9278; found: 2583.9256.
calcd for
1861.7313.
C
105H114O29Na [MþNaþ] 1861.7341; found:
3b-O-(2,3,4,6-Tetra-O-benzoyl-b-D-galactopyranosyl)-
(1!4)-2,3,6-tri-O-benzoyl-b-D-glucopyranosyl)oleanolic
acid, 2,3,4,6-tetra-O-benzoyl-b-D-glucopyranosyl-(1!6)-
2,3,4-tri-O-benzoyl-b-D-glucopyranosyl ester (4b)
3b-O-(2,3,4,6-Tetra-O-benzoyl-b-D-glucopyranosyl)-(1!4)-
2,3,6-tri-O-benzoyl-b-D-glucopyranosyl)oleanolic acid,
2,3,4,6-tetra-O-benzoyl-b-D-glucopyranosyl-(1!6)-2,3,4-
tri-O-benzoyl-b-D-glucopyranosyl ester (3b)
23
Yield: 61%; [a]D þ 23.1 (c 1.41, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.12–8.06 (m, 70 H, Ph-H), 5.84 (t, J ¼ 9.6 Hz, 1 H, H-
3000), 5.81 (d, J ¼ 8.0 Hz, 1H, H-1000), 5.78 (t, J ¼ 9.6 Hz, 1 H, H-30000),
5.76 (t, J ¼ 9.2 Hz, 1 H, H-30), 5.71 (dd, J ¼ 9.6, 3.7 Hz, 1 H, H-300),
5.58 (t, J ¼ 4.4 Hz, 1 H, H-400), 5.54 (dd, J ¼ 9.6, 8.0 Hz, 1 H, H-
2000), 5.52 (t, J ¼ 9.6 Hz, 1 H, H-40000), 5.50 (t, J ¼ 9.6 Hz, 1 H, H-4000),
5.48 (t, J ¼ 9.6 Hz, 1 H, H-40), 5.44 (dd, J ¼ 9.6, 7.9 Hz, 1 H, H-200),
5.29 (brs, 1 H, H-12), 5.05 (d, J ¼ 7.6 Hz, 1 H, H-10000), 4.88 (d,
J ¼ 7.6 Hz, 1 H, H-100), 4.70 (d, J ¼ 7.6 Hz, 1 H, H-10), 4.53–4.58
(m, 2 H, H-20, H-20000), 4.49 (dd, J ¼ 13.6, 5.4 Hz, 1 H, H-6000-1), 4.42
(dd, J ¼ 12.0, 4.8 Hz, 1 H, H-6000-2), 4.13–4.22 (m, 2 H, H-600-1, H-
60000-1), 3.98–4.05 (m, 2H, H-5000, H-600-2), 3.91–3.96 (m, 4 H, H-500,
H-50000, H-600-1, H-60000-1), 3.72–3.75 (m, 2 H, H-50, H-60-2), 2.97
(dd, J ¼ 11.2, 4.0 Hz, 1 H, H-3), 2.87 (dd, J ¼ 12.8, 2.8 Hz, 1 H, H-
18), 0.91, 0.86, 0.86, 0.74, 0.60, 0.58 0.40 (s each, 3 H each,
CH3 ꢂ 7); 13C NMR (CDCl3, 100 MHz): d 175.4 (C-28), 165.9,
165.7, 165.4, 165.3, 165.2, 165.1, 165.0, 164.9, 164.7, 164.4,
142.7 (C-13), 133.4, 133.2, 133.1, 133.0, 132.9, 132.8, 129.9,
129.8, 129.7, 129.6, 129.5, 129.4, 129.3, 129.2, 129.1, 128.8,
128.6, 128.5, 128.4, 128.3, 128.2, 128.1, 128.0, 122.6 (C-12),
103.0 (C-10), 100.8 (C-100), 99.9 (C-10000), 91.7 (C-1000), 90.6 (C-3),
75.0, 72.9, 72.8, 72.6, 72.5, 72.0, 71.8, 71.6, 71.5, 71.2, 70.6,
70.1, 69.8, 69.6, 69.1, 66.3, 65.1, 62.9, 62.6, 61.0, 55.1, 47.2,
46.6, 45.6, 42.2, 41.3, 40.8, 38.6, 38.4, 38.0, 36.3, 33.6, 32.8,
31.7, 31.4, 30.4, 29.5, 27.7, 27.4, 25.5, 25.2, 23.4, 23.1, 22.4,
20.6, 17.8, 16.2, 16.0, 15.0; HRMALDIMS: m/z calcd for
23
Yield: 63%; [a]D þ 50.6 (c 1.54, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.18–8.09 (m, 70 H, Ph-H), 5.85 (t, J ¼ 9.7 Hz, 1 H, H-
3000), 5.82 (t, J ¼ 9.6 Hz, 1 H, H-30000), 5.78 (d, J ¼ 7.9 Hz, 1H, H-1000),
5.75 (t, J ¼ 9.6 Hz, 1 H, H-30), 5.72 (t, J ¼ 9.6 Hz, 1 H, H-300), 5.57
(t, J ¼ 9.7 Hz, 1 H, H-400), 5.54 (dd, J ¼ 9.6, 7.9 Hz, 1 H, H-2000),
5.52 (t, J ¼ 9.6 Hz, 1 H, H-4000), 5.50 (t, J ¼ 9.6 Hz, 1 H, H-40000), 5.48
(t, J ¼ 9.6 Hz, 1 H, H-40), 5.39 (dd, J ¼ 9.6, 7.9 Hz, 1 H, H-200), 5.32
(t, J ¼ 3.6 Hz, 1 H, H-12), 5.06 (d, J ¼ 7.7 Hz, 1 H, H-10000), 4.93 (d,
J ¼ 7.9 Hz, 1 H, H-100), 4.85 (d, J ¼ 7.8 Hz, 1 H, H-10), 4.59 (dd,
J ¼ 9.6, 7.8 Hz, 1 H, H-20), 4.54 (dd, J ¼ 9.6, 7.7 Hz, 1 H, H-20000),
4.48 (dd, J ¼ 12.3, 3.9 Hz, 1 H, H-6000-1), 4.42 (dd, J ¼ 12.3, 5.3 Hz,
1 H, H-6000-2), 4.27 (dd, J ¼ 12.1, 3.3 Hz, 1 H, H-600-1), 4.19 (dd,
J ¼ 12.3, 4.8 Hz, 1 H, H-60000-1), 4.08 (dd, J ¼ 12.1, 4.7 Hz, 1 H, H-
600-2), 3.96–4.03 (m, 2 H, H-500, H-5000), 3.93 (m, 1 H, H-50000), 3.83–
3.87 (m, 2 H, H-50, H-60-1), 3.73–3.79 (m, 2 H, H-60-2, H-60000-2),
2.98 (dd, J ¼ 11.7, 4.1 Hz, 1 H, H-3), 2.89 (dd, J ¼ 13.7, 3.7 Hz, 1
H, H-18), 0.90, 0.85, 0.81, 0.75, 0.63, 0.59, 0.43 (s each, 3 H each,
CH3 ꢂ 7); 13C NMR (CDCl3, 100 MHz): d 175.5 (C-28), 165.9,
165.7, 165.5, 165.3, 165.2, 165.1, 165.0, 164.9, 164.7, 164.5,
142.8 (C-13), 133.3, 133.2, 133.1, 133.0, 129.9, 129.8, 129.7,
129.6, 129.5, 129.4, 129.3, 129.2, 129.0, 128.9, 128.6, 128.5,
128.4, 128.3, 128.2, 128.1, 128.0, 122.7 (C-12), 103.0 (C-10),
100.9 (C-100), 100.1 (C-10000), 91.9 (C-1000), 90.9 (C-3), 75.2, 73.1,
72.9, 72.8, 72.7, 72.6, 72.3, 72.2, 72.1, 71.9, 71.8, 70.2, 69.8,
69.4, 69.3, 64.4, 63.1, 62.6, 55.2, 47.3, 46.7, 45.7, 41.4, 40.7,
C
152H144O37Na [MþNaþ] 2583.9277; found: 2583.9293.
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T. Guo et al.
Archiv der Pharmazie
3b-O-(2,3,4-Tri-O-benzoyl-b-D-xylopyranosyl)oleanolic
acid, 2,3,4,6-tetra-O-benzoyl-b-D-glucopyranosyl ester (5b)
4.34 (ddd, J ¼ 3.7, 8.2, 10.1 Hz, 1H, H-50), 3.96 (d, J ¼ 10.5 Hz,
1H, H-600-1), 3.88 (dd, J ¼ 4.6, 11.5 Hz, 1H, H-600-2), 2.94 (dd,
J ¼ 10.1, 6.0 Hz, 1H, H-3), 2.70 (dd, J ¼ 13.7, 3.7Hz, 1H, H-18),
0.90, 0.85, 0.81, 0.78, 0.73, 0.66, 0.38 (s each, 3H each, CH3 ꢂ 7);
13C NMR (CDCl3, 150 MHz): d 175.6 (C-28), 166.0, 165.6, 165.5,
165.3, 165.1, 164.6, 142.9 (C-13), 133.4, 133.2, 133.1, 133.0,
129.9, 129.7, 129.4, 128.7, 128.4, 128.3, 122.8 (C-12), 100.1 (C-
10), 91.9 (C-100), 78.9 (C-3), 75.2, 72.9, 72.7, 71.8, 70.3, 69.8, 69.3,
66.5, 63.1, 55.1, 47.5, 46.8, 45.7, 41.5, 40.9, 38.9, 38.6, 38.4,
36.9, 33.7, 32.9, 31.9, 31.7, 30.5, 29.7, 28.1, 27.9, 27.2, 25.4,
23.6, 23.4, 22.6, 18.2, 16.5, 15.6, 15.3; HRESIMS: m/z calcd for
23
Yield: 68%; [a]D þ 16.3 (c 1.11, CHCl3); 1H NMR (CDCl3,
400 MHz): d 7.33–8.03 (m, 35H, Ph-H), 5.97 (t, J ¼ 9.6 Hz, 1H, H-
300), 5.93 (d, J ¼ 8.7 Hz, 1H, H-100), 5.78 (t, J ¼ 8.5 Hz, 1H, H-30),
5.71–5.74 (m, 2H, H-200, H-400), 5.44 (dd, J ¼ 8.2, 6.4 Hz, 1H, H-20),
5.30 (dt, J ¼ 8.7, 4.6 Hz, 1H, H-40), 5.27 (t, J ¼ 3.7 Hz, 1H, H-12),
4.81 (d, J ¼ 6.4 Hz, 1H, H-10), 4.53 (dd, J ¼ 11.5, 2.8 Hz, 1H, H-600-
1), 4.47 (dd, J ¼ 11.0, 5.0 Hz, 1H, H-600-2), 4.43 (dd, J ¼ 11.9,
4.6 Hz, 1H, H-50-1), 4.24-4.26 (m, 1H, H-500), 3.61 (dd, J ¼ 11.9,
7.8 Hz, 1H, H-50-2), 3.10 (dd, J ¼ 11.9, 4.6 Hz, 1H, H-3), 2.80 (dd,
J ¼ 13.3, 3.7 Hz, 1H, H-18), 0.93, 0.84, 0.81, 0.74, 0.72, 0.62, 0.42
(s each, 3 H each, CH3 ꢂ 7); 13C NMR (CDCl3, 100 MHz): d 175.3
(C-28), 165.8, 165.3, 165.2, 165.0, 164.9, 164.7, 142.3 (C-13),
133.7, 133.5, 133.1, 133.0, 132.9, 132.8, 129.9, 129.7, 129.5,
129.4, 129.3, 129.0, 128.8, 128.5, 128.1, 128.0, 122.4 (C-12),
103.1 (C-10), 91.9 (C-100), 90.3 (C-3), 75.3, 72.7, 72.3, 71.8, 71.3,
70.7, 70.3, 69.3, 65.5, 55.6, 47.3, 46.7, 45.6, 42.3, 41.3, 40.8,
38.7, 38.4, 38.1, 36.3, 33.5, 32.8, 31.7, 31.5, 30.4, 29.7, 27.3,
27.1, 25.6, 25.2, 23.4, 23.1, 22.4, 20.7, 17.8, 16.3, 16.1, 15.0;
HRESIMS: m/z calcd for C90H94O19Na [MþNaþ] 1501.6281;
found: 1501.6299.
C
91H97O20 [MþHþ] 1509.6567; found: 1509.6521.
Oleanolic acid, 28-O-2,3,4-tri-O-benzoyl-a-L-
rhamnopyranosyl-(1!4)-2,3,4,6-tetra-O-benzoyl-b-D-
glucopyranosyl-(1!6)-2,3,4-tri-O-benzoyl-b-D-
glucopyranosyl ester (8b)
23
Yield: 87%; [a]D þ 48.3 (c 1.10, CHCl3); 1H NMR (CDCl3,
400MHz): d 7.26–8.05 (m, 45 H, Ph-H), 5.87 (t, J ¼ 9.7 Hz, 1 H,
H-30), 5.85 (d, J ¼ 8.2 Hz, 1 H, H-10), 5.82 (t, J ¼ 9.7 Hz, 1 H, H-300),
5.73 (dd, J ¼ 9.9, 3.3 Hz, 1 H, H-3000), 5.63 (dd, J ¼ 9.6, 8.2 Hz, 1 H,
H-20), 5.56 (br s, 1 H, H-2000), 5.53 (t, J ¼ 9.7 Hz, 1 H, H-40), 5.47 (t,
J ¼ 9.9 Hz, 1 H, H-4000), 5.44 (dd, J ¼ 9.7, 7.8 Hz, 1 H, H-200), 5.39 (br
s, 1 H, H-12), 5.27 (dd, J ¼ 12.5, 3.7 Hz, H-60-1), 5.23 (s, 1 H, H-1000),
5.01 (d, J ¼ 7.8 Hz, 1 H, H-100), 4.99 (dd, J ¼ 12.1, 3.2 Hz, H-600-1),
4.67 (dd, J ¼ 12.1, 3.9 Hz, 1 H, H-600-2), 4.29 (t, J ¼ 9.7 Hz, 1 H, H-
400), 4.06 (m, 1 H, H-5000), 4.01 (dd, J ¼ 12.5, 5.1 Hz, 1 H, H-60-2),
3.91–3.96 (m, 2 H, H-50, H-500), 3.21 (dd, J ¼ 11.9, 3.7 Hz, 1 H, H-3),
2.95(dd,J ¼ 14.3,3.7 Hz,1H,H-18),0.76(d,J ¼ 6.0 Hz,3H,H-6000),
0.99, 0.96, 0.87, 0.85, 0.81, 0.71, 0.50 (s each, 3 H each, CH3 ꢂ 7);
13C NMR (CDCl3, 100MHz): d 175.6 (C-28), 165.9, 165.6, 165.4,
165.2, 165.1, 143.1 (C-13), 133.3, 133.2, 133.0, 130.1, 130.0,
129.9,129.7,129.5,128.9,128.4,128.0,122.7(C-12),100.1(C-100),
99.0 (C-1000), 91.9 (C-10), 89.3 (C-3), 75.3, 74.0, 73.9, 73.6, 73.5,
73.4, 73.2, 73.0, 72.6, 72.4, 71.4, 70.5, 69.5, 67.7, 55.3, 46.9, 41.6,
39.3, 38.9, 37.9, 33.0, 30.6, 29.7, 27.9, 25.3, 23.6, 17.1, 16.4, 15.1;
HR-MALDI-MS: m/z calcd for C111H115O26 [MþHþ] 1863.7671;
found: 1863.7693.
Typical procedure for the synthesis of compounds 6b–8b
To a solution of oleanolic acid (240 mg, 0.53 mmol) and
bromide 7a, 8a, or 9a (0.68 mmol, 1.2 equiv.) in CH2Cl2 (8 mL)
were added K2CO3 (187 mg, 1.33 mmol), water (5.0 mL), and
Bu4NBr (69 mg, 0.21 mmol). The resulting mixture was refluxed
for 6 h, and then diluted with CH2Cl2. The organic phase, after
being washed with water and brine, was dried with Na2SO4,
and then concentrated in vacuo. The residue was purified by a
silica gel column chromatography (5:1 to 3:1, petroleum ether/
EtOAc) to afford the product 7, 8, or 9 as a white solid.
Oleanolic acid, 28-O-2,3,4,6-tetra-O-benzoyl-b-D-
glucopyranosyl ester (6b)
23
Yield: 89%; [a]D þ 93.3 (c 2.50, CHCl3); 1H NMR (DMSO-d6,
600 MHz): d 7.36–7.81 (m, 20 H, Ph-H), 6.15 (d, J ¼ 8.1 Hz, 1H, H-
10), 6.09 (t, J ¼ 9.7 Hz, 1H, H-30), 5.75 (s, 1 H, C3-OH), 5.43 (t,
J ¼ 9.7 Hz, 1 H, H-40), 5.37 (dd, J ¼ 9.7, 8.1 Hz, 1 H, H-20), 5.19 (t,
J ¼ 3.7 Hz, 1 H, H-12), 4.49 (dd, J ¼ 12.3, 3.6 Hz, 1 H, H-60-1), 4.45
(dd, J ¼ 12.3, 5.0 Hz, 1 H, H-60-2), 4.31 (m, 1 H, H-50), 2.97 (dd,
J ¼ 13.7, 4.3 Hz, 1 H, H-3), 2.81 (dd, J ¼ 13.7, 3.7 Hz, 1H, H-18),
0.93, 0.85, 0.81, 0.77, 0.73, 0.67, 0.43 (s each, 3 H each,
CH3 ꢂ 7); HRESIMS: m/z calcd for C64H75O12 [MþHþ]
1035.5253; found: 1035.5277.
Typical procedure for the synthesis of compounds 9b–16b
A mixture of compound 17 (100 mg, 0.11 mmol), trichloro-
acetimidate 3a–6a, or 10a–13a (0.14 mmol, 1.2 equiv.) and
̊
powdered 4 A molecular sieves in dry CH2Cl2 (5 mL) were
stirred for 30 min at room temperature and then cooled to 0°
C. TMSOTf (15 mL, 0.09 mmol, 0.3 equiv.) was added slowly.
After being stirred at ꢁ78°C for 30 min, the reaction mixture
was warmed up to room temperature for 30 min. The reaction
was quenched by addition of Et3N and then filtered. The
filtrate was concentrated and purified by a silica gel column
chromatography (petroleum ether/EtOAc) to afford the
products 9b–16b.
Oleanolic acid, 28-O-2,3,4,6-tetra-O-benzoyl-b-D-
glucopyranosyl-(1!6)-2,3,4-tri-O-benzoyl-b-D-
glucopyranosyl ester (7b)
23
Yield: 85%; [a]D þ 37.1 (c 1.65, CHCl3); 1H NMR (DMSO-d6,
600 MHz): d 7.36–7.97 (m, 35 H, Ph-H), 6.14 (d, J ¼ 8.3 Hz, 1H, H-
10), 6.10 (t-like, J ¼ 9.7, 9.6 Hz, 1H, H-30), 5.88 (t-like, J ¼ 9.7,
9.6 Hz, 1H, H-300), 5.77 (s, 1 H, C3-OH), 5.39–5.45 (m, 3 H, H-20, H-
200, H-40), 5.20 (d, J ¼ 7.8 Hz, 1H, H-100), 5.17 (s, 1 H, H-12), 4.50
(ddd, J ¼ 4.6, 6.9, 10.1 Hz, 1H, H-500), 4.46 (m, 2H, H-60-1, H-60-2),
3-O-2,3,4,6-Tetra-O-benzoyl-b-D-glucopyranoside
oleanolic acid (9b)
23
1
Yield: 88%; [a]D ¼ þ 65.3 (c 1.56, CHCl3); H NMR (600 MHz,
DMSO-d6): d 12.03 (s, 1H, COOH), 7.43–7.85 (m, 20H, Ph-H),
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Oleanolic Acid Saponins as a-Glucosidase Inhibitors
Archiv der Pharmazie
5.99 (t, J ¼ 9.4 Hz, 1H, H-40), 5.53 (t, J ¼ 9.4 Hz, 1H, H-30), 5.38
(dd, J ¼ 9.4, 8.2 Hz, 1H, H-20), 5.17 (d, J ¼ 8.2 Hz, 1H, H-10), 5.16
(br s, 1H, H-12), 4.56 (dd, J ¼ 12.1, 7.1 Hz, 1H, H-60-1), 4.43–4.48
(m, 2H, H-50, H-60-2), 3.12 (dd, J ¼ 12.1, 4.4 Hz, 1H, H-3), 2.74
(dd, J ¼ 13.7, 3.9 Hz, 1H, H-18), 1.06, 0.87, 0.87, 0.76, 0.65, 0.64,
0.51 (s each, 3H each, CH3 ꢂ 7); 13C NMR (150 MHz, CDCl3): d
184.0 (C-28), 166.2, 166.1, 165.3, 165.2, 143.8 (C-13), 133.5,
133.4, 133.3, 130.1, 130.0, 129.9, 129.1, 129.0, 128.8, 128.6,
128.5, 128.4, 122.8 (C-12), 103.5 (C-10), 91.0 (C-3), 73.2, 72.3,
72.2, 70.5, 63.6, 60.6, 55.6, 47.8, 46.7, 46.1, 41.7, 41.1, 39.4,
38.9, 38.4, 36.8, 34.0, 33.3, 32.7, 32.6, 30.9, 27.8, 26.1, 26.0,
23.8, 23.5, 23.1, 18.2, 17.2, 16.3, 15.3, 14.4; HRESIMS: m/z calcd
for C64H74O12Na [MþNa]þ 1057.5093; found: 1057.5078.
H-20), 5.47 (t, J ¼ 9.9 Hz, 1H, H-40), 5.12 (t, J ¼ 3.6 Hz, 1H, H-12),
5.01 (s, 1H, H-10), 4.19 (m, 1H, H-50), 3.23 (dd, J ¼ 12.1, 3.7 Hz,
1H, H-3), 2.78 (dd, J ¼ 13.7, 3.7 Hz, 1H, H-18), 1.19 (d, J ¼ 6.0 Hz,
3H, H-60), 1.05, 0.99, 0.89, 0.86, 0.71, 0.68, 0.53 (s each, 3H
each, CH3 ꢂ 7); HRESIMS: m/z calcd for C57H71O10 [MþH]þ
915.5041; found: 915.5067.
3-O-2,3,4-Tri-O-benzoyl-a-L-fucopyranoside oleanolic acid
(14b)
25
1
Yield: 83%; [a]D ¼ ꢁ 27.9 (c 1.90, CHCl3); H NMR (600 MHz,
DMSO-d6): d 12.05 (s, 1H, COOH), 7.34–7.99 (m, 15H, Ph-H),
5.71 (dd, J ¼ 10.1, 3.2 Hz, 1H, H-30), 5.59 (d, J ¼ 4.1 Hz, 1H, H-40),
5.44 (dd, J ¼ 10.6, 8.3 Hz, 1H, H-20), 5.14 (t-like, J ¼ 3.7, 3.2 Hz,
1H, H-12), 5.07 (d, J ¼ 8.3 Hz, 1H, H-10), 4.29 (dd, J ¼ 13.3,
6.4 Hz, 1H, H-50), 3.27 (dd, J ¼ 11.4, 4.1 Hz, 1H, H-3), 2.72 (dd,
J ¼ 13.7, 4.1 Hz, 1H, H-18), 1.20 (d, J ¼ 6.4 Hz, 3H, H-60), 1.09,
0.99, 0.87, 0.87, 0.70, 0.68, 0.58 (s each, 3H each, CH3 ꢂ 7);
HRESIMS: m/z calcd for C57H70O10Na [MþNa]þ 937.4867;
found: 937.4835.
3-O-2,3,4-Tri-O-benzoyl-b-D-xylopyranoside oleanolic acid
(10b)
23
1
Yield: 87%; [a]D ¼ þ 78.1 (c 1.78, CHCl3); H NMR (600 MHz,
DMSO-d6): d 12.01 (s, 1H, COOH), 7.41–7.81 (m, 15H, Ph-H),
5.69 (t, J ¼ 9.7 Hz, 1H, H-30), 5.56 (m, 1H, H-40), 5.43 (dd, J ¼ 9.7,
7.6 Hz, 1H, H-20), 5.19 (br s, 1H, H-12), 5.15 (d, J ¼ 7.6 Hz, 1H, H-
10), 4.73 (dd, J ¼ 12.3, 4.5 Hz, 1H, H-50-1), 4.57 (dd, J ¼ 12.3,
6.9 Hz, 1H, H-50-2), 3.10 (dd, J ¼ 11.9, 3.7 Hz, 1H, H-3), 2.79 (dd,
J ¼ 13.7, 4.3 Hz, 1H, H-18), 1.06, 0.89, 0.87, 0.76, 0.65, 0.63, 0.49
(s each, 3H each, CH3 ꢂ 7); HRESIMS: m/z calcd for C56H69O10
[MþH]þ 901.4885; found: 901.4867.
3-O-2,3,4,6-Tetra-O-benzoyl-b-D-glucopyranosyl-(1!4)-b-
D-glucopyranoside oleanolic acid (15b)
25
Yield: 85%; [a]D ¼ þ 81.67 (c 1.20, CHCl3); 1H NMR (600 MHz,
DMSO-d6): d 12.05 (s, 1H, COOH), 7.29–8.03 (m, 35H, Ph-H),
5.77 (t, 1H, J ¼ 9.6 Hz, H-300), 5.69 (t, 1H, J ¼ 9.6 Hz, H-30), 5.63 (t,
1H, J ¼ 9.6 Hz, H-400), 5.50 (dd, J ¼ 9.6, 7.8 Hz, 1H, H-200), 5.35 (d,
J ¼ 7.8 Hz, 1H, H-100), 5.21 (dd, J ¼ 9.7, 8.0 Hz, 1H, H-20), 5.13 (t,
J ¼ 3.6 Hz, 1H, H-12), 4.98 (d, J ¼ 8.0 Hz, 1H, H-10), 4.73 (dd,
J ¼ 12.1, 4.3 Hz, 1H, H-60-1), 4.65 (m, 1H, H-500), 4.52 (dd,
J ¼ 12.1, 6.7 Hz, 1H, H-60-2), 4.47 (t, J ¼ 9.6 Hz, 1H, H-40), 3.96
(dd, J ¼ 11.9, 5.1 Hz, 1H, H-600-1), 3.89 (m, 1H, H-50), 3.67 (dd,
J ¼ 11.9, 6.4 Hz, 1H, H-600-2), 3.06 (dd, J ¼ 11.7, 3.7 Hz, 1H, H-3),
2.77 (dd, J ¼ 13.7, 3.7 Hz, 1H, H-18), 1.03, 0.89, 0.86, 0.71, 0.65
0.58, 0.53 (s each, 3 H each, CH3 ꢂ 7); 13C NMR (100 MHz,
CDCl3): d 183.5 (C-28), 166.1, 165.5, 165.4, 165.1, 143.5 (C-13),
133.9, 133.6, 133.1, 130.0, 129.9, 129.7, 128.9, 128.6, 128.5,
128.3, 122.5 (C-12), 103.7 (C-10), 100.9 (C-100), 90.7 (C-3), 76.3,
73.9, 73.0, 72.3, 71.9, 71.5, 70.1, 69.8, 62.9, 61.5, 55.7, 47.7,
46.9, 46.1, 41.7, 41.3, 39.4, 38.8, 38.5, 36.8, 33.9, 33.6, 32.6,
30.9, 29.9, 27.8, 26.1, 25.7, 23.8, 23.5, 18.1, 17.2, 16.3, 15.1;
HRMALDIMS: m/z calcd for C91H96O20Na [MþNa]þ 1531.6379;
found: 1531.6391.
3-O-2,3,4,6-Tetra-O-benzoyl-b-D-galactopyranoside
oleanolic acid (11b)
23
1
Yield: 89%; [a]D ¼ þ 98.1 (c 2.01, CHCl3); H NMR (600 MHz,
DMSO-d6): d 12.05 (s, 1H, COOH), 7.43–7.87 (m, 20H, Ph-H),
5.96 (t, J ¼ 3.7 Hz, 1H, H-40), 5.51 (dd, J ¼ 9.7, 3.7 Hz, 1H, H-30),
5.33 (dd, J ¼ 9.7, 8.0 Hz, 1H, H-20), 5.21 (d, J ¼ 8.0 Hz, 1H, H-10),
5.17 (br s, 1H, H-12), 4.56 (dd, J ¼ 12.5, 3.9 Hz, 1H, H-60-1), 4.46
(m, 1H, H-50), 4.39 (dd, J ¼ 12.5, 5.3 Hz, 1H, H-60-2), 3.12 (dd,
J ¼ 12.3, 4.4 Hz, 1H, H-3), 2.78 (dd, J ¼ 13.7, 4.3 Hz, 1H, H-18),
1.01, 0.93, 0.85, 0.71, 0.65, 0.64, 0.53 (s each, 3H each, CH3 ꢂ 7);
HRESIMS: m/z calcd for C64H75O12 [MþH]þ 1035.5253; found:
1035.5229.
3-O-2,3,4,6-Tetra-O-benzoyl-a-D-mannopyranoside
oleanolic acid (12b)
23
1
Yield: 81%; [a]D ¼ þ 49.6 (c 1.21, CHCl3); H NMR (600 MHz,
DMSO-d6): d 12.05 (s, 1H, COOH), 7.37–8.42 (m, 20H, Ph-H),
5.97 (t, J ¼ 10.1 Hz, 1H, H-40), 5.73 (dd, J ¼ 10.1, 3.2 Hz, 1H, H-
30), 5.53 (dd, J ¼ 3.2, 1.9 Hz, 1H, H-20), 5.37 (s, 1H, H-10), 5.17
(br s, 1H, H-12), 4.66 (m, 1H, H-60-1), 4.49–4.54 (m, 2H, H-50, H-
60-2), 3.31 (m, 1H, H-3), 2.74 (dd, J ¼ 13.7, 4.1 Hz, 1H, H-18),
1.11, 1.09, 0.93, 0.91, 0.88, 0.88, 0.74 (s each, 3H each, CH3 ꢂ 7);
HRESIMS: m/z calcd for C64H74O12Na [MþNa]þ 1057.5078;
found: 1057.5071.
3-O-2,3,4,6-Tetra-O-benzoyl-b-D-galactopyranosyl-(1!4)-
b-D-glucopyranoside oleanolic acid (16b)
25
Yield: 83%; [a]D ¼ þ 100.7 (c 1.51, CHCl3); 1H NMR (600 MHz,
DMSO-d6): d 12.04 (s, 1H, COOH), 7.22–7.97 (m, 35H, Ph-H),
5.75–5.79 (m, 2H, H-30, H-300), 5.67 (d, 1H, J ¼ 3.2 Hz, H-400), 5.43
(dd, J ¼ 7.8, 10.1 Hz, 1H, H-200), 5.31 (d, J ¼ 7.8 Hz, 1H, H-100),
5.24 (dd, J ¼ 9.7, 8.2 Hz, 1H, H-20), 5.16 (br s, 1H, H-12), 5.00 (d,
J ¼ 8.2 Hz, 1H, H-10), 4.62 (dd, J ¼ 11.9, 8.2 Hz, 1H, H-60-1), 4.52
(d, J ¼ 10.6 Hz, 1H, H-60-2), 4.43 (t-like, J ¼ 6.9, 6.8 Hz, 1H, H-500),
4.31 (t, J ¼ 9.6 Hz, 1H, H-40), 4.09 (t, J ¼ 9.6 Hz, 1H, H-50), 3.82
(dd, J ¼ 11.5, 6.9 Hz, 1H, H-600-1), 3.60 (dd, J ¼ 11.0, 6.4 Hz, 1H,
H-600-2), 3.00 (dd, J ¼ 11.9, 4.1 Hz, 1H, H-3), 2.73 (dd, J ¼ 13.3,
3.7 Hz, 1H, H-18), 1.05, 0.89, 0.88, 0.71, 0.64 0.58, 0.47 (s each,
3-O-2,3,4-Tri-O-benzoyl-a-L-rhamnopyranoside oleanolic
acid (13b)
25
Yield: 89%; [a]D ¼ 13.5 (c 1.15, CHCl3); 1H NMR (600 MHz,
DMSO-d6): d 12.01 (s, 1H, COOH), 7.29–7.87 (m, 15H, Ph-H),
5.79 (dd, J ¼ 9.9, 3.5 Hz, 1H, H-30), 5.63 (dd, J ¼ 3.7, 1.9 Hz, 1H,
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T. Guo et al.
Archiv der Pharmazie
3H each, CH3 ꢂ 7); 13C NMR (150 MHz, CDCl3): d 183.8 (C-28),
166.0, 165.8, 165.4, 165.3, 143.8 (C-13), 133.8, 133.6, 133.5,
130.0, 129.9, 129.8, 129.7, 128.8, 128.6, 128.5, 128.4, 122.8 (C-
12), 103.4 (C-10), 101.3 (C-100), 90.9 (C-3), 73.2, 73.0, 72.1, 71.9,
71.6, 70.1, 67.8, 62.9, 61.4, 55.6, 47.7, 46.7, 46.1, 41.7, 41.1,
39.4, 38.8, 38.4, 36.8, 33.9, 33.3, 32.6, 30.8, 29.9, 27.8, 26.1,
25.9, 23.8, 23.0, 18.1, 17.2, 16.3, 15.3; HRMALDIMS: m/z calcd
for C91H96O20Na [MþNa]þ 1531.6379; found: 1531.6387.
100 MHz): d 178.1 (C-28), 144.9 (C-13), 123.9 (C-12), 106.6 (C-
10), 105.1 (C-100), 95.7 (C-1000), 90.9 (C-3), 80.6, 78.7, 78.3, 77.0,
76.6, 76.2, 75.3, 74.8, 73.9, 72.5, 71.1, 70.5, 62.6, 62.4, 62.0,
57.1, 48.0, 47.3, 43.0, 42.6, 40.7, 40.2, 39.9, 37.9, 34.0, 33.5,
33.2, 31.6, 28.9, 28.5, 27.3, 26.3, 24.6, 24.1, 19.5, 17.8, 17.1,
16.3; HRESIMS: m/z calcd for C48H79O18 [MþHþ] 943.5261;
found: 943.5249.
3b-O-(b-D-Glucopyranosyl)-(1!4)-b-D-glucopyranosyl)-
oleanolic acid, b-D-glucopyranosyl-(1!6)-b-D-
glucopyranosyl ester (3)
Typical procedure for the synthesis of compounds 1–16
To a solution of fully protected oleanolic acid saponins 1b–16b
(100 mg) in dry 1:2 CH2Cl2/MeOH (20 mL) was added a newly
prepared NaOMe in MeOH solution (1.0 mol/L, 0.20 mL). The
mixture was stirred at rt for 5 h and neutralized with Dowex
23
Yield: 89%; [a]D þ 3.4 (c 0.87, CH3OH); 1H NMR (CD3OD,
400 MHz): d 5.32 (d, J ¼ 8.0 Hz, 1 H, H-1000), 5.19 (brs, 1 H, H-12),
4.37 (d, J ¼ 7.6 Hz, 1 H, H-10), 4.30 (d, J ¼ 7.6 Hz, 1 H, H-100), 4.28
Hþ resin to pH
7
and then filtered. The filtrate was
(d, J ¼ 7.6 Hz, 1 H, H-10000), 4.08 (dd, J ¼ 11.2, 2.1 Hz, 1 H, H-60000
-
concentrated and the resulting residue was subjected to a
silica gel column chromatography (MeOH/CHCl3) to give the
target compounds 1–16 as white amorphous solids.
1), 3.84 (dd, J ¼ 12.0, 3.3 Hz, 1 H, H-600-1), 3.82 (dd, J ¼ 12.3,
4.3 Hz, 1 H, H-60-1), 3.80 (dd, J ¼ 12.0, 4.9 Hz, 1 H, H-600-2), 3.78
(dd, J ¼ 11.9, 3.1 Hz, 1 H, H-6000-1), 3.72 (dd, J ¼ 11.6, 4.0 Hz, 1 H,
H-60000-2), 3.63 (t, J ¼ 10.0 Hz, 1 H, H-300), 3.60 (dd, J ¼ 11.9,
4.9 Hz, 1 H, H-6000-2), 3.57 (t, J ¼ 9.7 Hz, 1 H, H-3000), 3.51 (t,
J ¼ 9.2 Hz, 1 H, H-4000), 3.47 (t, J ¼ 9.2 Hz, 1 H, H-2000), 3.45 (dd,
J ¼ 9.6, 7.6 Hz, 1 H, H-200), 3.43 (dd, J ¼ 10.1, 7.7 Hz, 1 H, H-20000),
3.37–3.40 (m, 2 H, H-20, H-5000), 3.37 (t, J ¼ 9.7 Hz, 1 H, H-40000),
3.34 (t, J ¼ 10.0 Hz, 1 H, H-40), 3.22 (m, 1 H, H-500), 3.17–3.21 (m,
2 H, H-50, H-50000), 3.15 (t, J ¼ 9.6 Hz, 1 H, H-400), 3.10 (dd, J ¼ 13.7,
3.6 Hz, 1 H, H-3), 2.80 (dd, J ¼ 13.2, 2.8 Hz, 1 H, H-18), 1.10,
3b-O-(b-D-Glucopyranosyl)-(1!4)-b-D-glucopyranosyl)
oleanolic acid, b-D-glucopyranosyl ester (1)
23
Yield: 91%; [a]D þ 11.7 (c 0.98, CH3OH); 1H NMR (CD3OD,
400 MHz): d 5.32 (d, J ¼ 8.0 Hz, 1H, H-1000), 5.19 (brs, 1H, H-12),
4.35 (d, J ¼ 8.0 Hz, 1H, H-10), 4.28 (d, J ¼ 8.0 Hz, 1H, H-100), 3.77–
3.83 (m, 3H, H-60-1, H-400, H-600-1), 3.74–3.76 (m, 2H, H-6000-1, H-
600-2), 3.63 (dd, J ¼ 12.5, 3.4 Hz, 1H, H-6000-2), 3.60 (t, J ¼ 9.4 Hz,
1H, H-4000), 3.58 (dd, J ¼ 11.9, 5.1 Hz, 1H, H-60-2), 3.50 (t,
J ¼ 9.2 Hz, 1H, H-3000), 3.43 (t, J ¼ 9.2 Hz, 1H, H-300), 3.32–3.37 (m,
4H, H-20, H-200, H-2000, H-40), 3.26–3.29 (m, 2H, H-30, H-5000), 3.11–
3.16 (m, H-50, H-500), 3.09 (dd, J ¼ 11.6, 4.0 Hz, 1H, H-3), 2.80
(dd, J ¼ 13.2, 3.2 Hz, 1H, H-18), 1.09, 0.99, 0.89, 0.87, 0.85, 0.79,
0.74 (s each, 3 H each, CH3 ꢂ 7); 13C NMR (CD3OD, 100 MHz): d
178.1 (C-28), 144.9 (C-13), 123.9 (C-12), 106.6 (C-10), 104.6 (C-
100), 95.8 (C-1000), 90.9 (C-3), 80.7, 78.8, 78.3, 78.2, 77.9, 76.6,
76.3, 75.4, 75.0, 74.0, 71.4, 71.1, 62.5, 62.0, 57.1, 48.1, 47.3,
43.0, 42.6, 40.8, 40.2, 39.9, 37.9, 34.9, 34.0, 33.5, 33.2, 31.6,
28.9, 28.5, 27.1, 26.3, 24.6, 24.0, 19.4, 17.8, 17.0, 16.1;
HRESIMS: m/z calcd for C48H79O18 [MþHþ] 943.5261; found:
943.5273.
0.99, 0.90, 0.88, 0.86, 0.79, 0.74 (s each, 3 H each, CH3 ꢂ 7); 13
C
NMR (CD3OD, 100 MHz): d 178.1 (C-28), 144.9 (C-13), 123.8 (C-
12), 106.6 (C-10), 104.7 (C-100), 104.6 (C-10000), 95.8 (C-1000), 90.9(C-
3), 80.7, 78.2, 78.0, 77.9, 76.6, 76.3, 75.4, 75.2, 75.0, 73.9, 71.6,
71.4, 70.9, 69.5, 62.8, 62.5, 62.0, 57.1, 43.0, 42.6, 40.8, 39.9,
37.9, 34.9, 34.0, 33.6, 33.2, 31.6, 29.0, 28.6, 27.1, 26.4, 24.6,
24.1, 19.4, 17.9, 17.1, 16.2; HRESIMS: m/z calcd for C54H89O23
[MþHþ] 1105.5789; found: 1105.5771.
3b-O-(b-D-Galactopyranosyl)-(1!4)-b-D-glucopyranosyl)-
oleanolic acid, b-D-glucopyranosyl-(1!6)-b-D-
glucopyranosyl ester (4)
23
Yield: 90%; [a]D þ 10.7 (c 0.90, CH3OH); 1H NMR (CD3OD,
400 MHz): d 5.35 (d, J ¼ 7.9 Hz, 1 H, H-1000), 5.21 (brs, 1 H, H-12),
4.38 (d, J ¼ 7.8 Hz, 1 H, H-10), 4.35 (d, J ¼ 7.6 Hz, 1 H, H-100), 4.30
3b-O-(b-D-Galactopyranosyl)-(1!4)-b-D-glucopyranosyl)-
oleanolic acid, b-D-glucopyranosyl ester (2)
(d, J ¼ 8.0 Hz, 1 H, H-10000), 4.06 (dd, J ¼ 12.3, 3.5 Hz, 1 H, H-60000
-
23
Yield: 92%; [a]D þ 5.7 (c 1.01, CH3OH); 1H NMR (CD3OD,
1), 3.82 (dd, J ¼ 12.1, 2.3 Hz, 1 H, H-600-1), 3.80 (dd, J ¼ 12.1,
3.9 Hz, 1 H, H-6000-1), 3.78 (dd, J ¼ 11.7, 3.1 Hz, 1 H, H-6000-1), 3.76
(dd, J ¼ 11.9, 3.6 Hz, 1 H, H-60-1), 3.72 (dd, J ¼ 12.3, 4.7 Hz, 1H,
H-60000-2), 3.65 (dd, J ¼ 12.1, 5.0 Hz, 1 H, H-6000-2), 3.63 (t,
J ¼ 9.9 Hz, 1 H, H-4000), 3.60 (t, J ¼ 9.9 Hz, 1 H, H-3000), 3.56 (t,
J ¼ 10.0 Hz, 1 H, H-300), 3.53 (dd, J ¼ 10.0, 7.8 Hz, 1 H, H-20000),
3.51 (dd, J ¼ 9.9, 7.9 Hz, 1 H, H-2000), 3.47 (dd, J ¼ 9.7, 7.6 Hz, 1 H,
H-20), 3.43 (dd, J ¼ 9.9, 7.8 Hz, 1 H, H-200), 3.40 (m, 1 H, H-5000),
3.37 (t, J ¼ 9.9 Hz, 1 H, H-40000), 3.35 (m, 1 H, H-50), 3.30 (t,
J ¼ 9.7 Hz, 1 H, H-40), 3.19–3.23 (m, 2 H, H-500, H-50000), 3.16 (t,
J ¼ 10.0 Hz, 1 H, H-400), 3.11 (dd, J ¼ 14.3, 3.7 Hz, 1 H, H-3), 2.79
(dd, J ¼ 13.7, 4.0 Hz, 1 H, H-18), 1.09, 0.99, 0.93, 0.89, 0.85, 0.79,
0.75 (s each, 3 H each, CH3 ꢂ 7); 13C NMR (CD3OD, 100 MHz): d
178.2 (C-28), 144.9 (C-13), 123.9 (C-12), 106.5 (C-10), 105.0 (C-
400 MHz): d 5.36 (d, J ¼ 8.1 Hz, 1 H, H-1000), 5.19 (brs, 1 H, H-12),
4.33 (d, J ¼ 8.0 Hz, 1 H, H-10), 4.29 (d, J ¼ 8.3 Hz, 1 H, H-100), 3.79
(dd, J ¼ 12.3, 3.7 Hz, 1 H, H-600-1), 3.77 (dd, J ¼ 12.1, 3.3 Hz, 1 H,
H-60-1), 3.74 (t, J ¼ 3.6 Hz, 1 H, H-400), 3.71 (dd, J ¼ 12.3, 3.3 Hz, 1
H, H-6000-1), 3.69 (dd, J ¼ 12.3, 4.9 Hz, 1 H, H-600-2), 3.64 (dd,
J ¼ 12.5, 3.7 Hz, 1 H, H-6000-2), 3.60 (t, J ¼ 9.7 Hz, 1 H, H-4000), 3.55
(dd, J ¼ 12.1, 5.3 Hz, 1 H, H-60-2), 3.51 (t, J ¼ 9.6 Hz, 1 H, H-3000),
3.47 (dd, J ¼ 9.6, 3.6 Hz, 1 H, H-300), 3.45 (dd, J ¼ 9.6, 8.3 Hz, 1 H,
H-200), 3.42 (dd, J ¼ 9.7, 8.1 Hz, 1 H, H-20), 3.35–3.39 (m, 2 H, H-
2000, H-40), 3.29 (t, J ¼ 9.6 Hz, 1 H, H-30), 3.25–3.28 (m, 1 H, H-500),
3.09–3.18 (m, 2H, H-50, H-5000), 3.07 (dd, J ¼ 11.7, 3.7 Hz, 1 H, H-
3), 2.81 (dd, J ¼ 13.7, 4.0 Hz, 1 H, H-18), 1.11, 0.98, 0.87, 0.79,
0.77, 0.74, 0.69 (s each, 3 H each, CH3 ꢂ 7); 13C NMR (CD3OD,
ß 2015 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim
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Oleanolic Acid Saponins as a-Glucosidase Inhibitors
Archiv der Pharmazie
100), 104.6 (C-10000), 95.8 (C-1000), 90.9 (C-3), 80.7, 78.1, 78.0, 77.8,
76.9, 76.2, 75.3, 75.1, 74.7, 73.5, 72.5, 71.5, 70.9, 70.5, 69.5,
62.6, 62.0, 57.1, 48.1, 47.3, 42.9, 42.6, 40.7, 39.9, 37.9, 34.9,
34.0, 33.5, 33.2, 31.6, 29.1, 28.5, 27.1, 26.3, 24.6, 24.3, 19.4,
17.6, 17.3, 16.1; HRESIMS: m/z calcd for C54H89O23 [MþHþ]
1105.5789; found: 1105.5801.
16.1; HRESIMS: m/z calcd for C42H69O13 [MþHþ] 781.4733;
found: 781.4750.
Oleanolic acid, 28-O-a-L-rhamnopyranosyl-(1!4)-b-D-
glucopyranosyl-(1!6)-b-D-glucopyranosyl ester (8)
Yield: 87%; 1H NMR (400 MHz, C5D5N) d: 6.25 (d, J ¼ 8.0 Hz, 1H,
H-100), 5.83 (br s, 1H, H-1000), 5.36 (t, J ¼ 3.7 Hz, 1H, H-12), 4.99 (d,
J ¼ 7.7 Hz, 1H, H-10), 4.93 (qd, J ¼ 9.3, 6.0 Hz, 1H, H-5000), 4.71
(dd, J ¼ 3.3, 1.7 Hz, 1H, H-2000), 4.63 (t, J ¼ 9.9 Hz, 1H, H-40), 4.56
(dd, J ¼ 9.6, 3.2 Hz, 1H, H-3000), 4.47 (t, J ¼ 9.6 Hz, 1H, H-300), 4.43
(t, J ¼ 9.6 Hz, 1H, H-4000), 4.36 (dd, J ¼ 12.5, 3.6 Hz, 1H, H-60-1),
4.31 (dd, J ¼ 9.6, 8.0 Hz, 1H, H-200), 4.29 (t, J ¼ 9.9 Hz, 1H, H-30),
4.23 (t, J ¼ 9.6 Hz, 1H, H-400), 4.21 (dd, J ¼ 12.5, 4.9 Hz, 1H, H-60-
2), 4.17 (dd, J ¼ 12.1, 3.3 Hz, 1H, H-600-1), 4.06–4.10 (m, 2 H, H-
50, H-500), 3.96 (dd, J ¼ 9.9, 7.7 Hz, 1H, H-20), 3.71 (dd, J ¼ 12.1,
5.0 Hz, 1H, H-600-2), 3.30 (dd, J ¼ 11.7, 3.7 Hz, 1H, H-3), 3.17 (dd,
J ¼ 13.7, 4.3 Hz, 1H, H-18), 1.68 (d, J ¼ 5.9 Hz, 3 H, H-6000), 1.27,
3b-O-(b-D-Xylopyranosyl)oleanolic acid, b-D-
glucopyranosyl ester (5)
23
Yield: 93%; [a]D þ 21.7 (c 1.30, CH3OH); 1H NMR (CD3OD,
600 MHz): d 5.38 (d, J ¼ 8.2 Hz, 1H, H-100), 5.25 (t, J ¼ 3.7 Hz, 1H,
H-12), 4.26 (t, J ¼ 7.3 Hz, 1H, H-10), 3.80–3.83 (m, 2H, H-400, H-50-
1), 3.68 (dd, J ¼ 12.4, 4.6 Hz, 1H, H-600-1), 3.43–3.48 (m, 1H, H-
40), 3.41 (t, J ¼ 9.1, 8.7 Hz, 1H, H-30), 3.33–3.36 (m, 3H, H-500, H-
600-2, H-50-2), 3.28 (t, J ¼ 9.1, 8.7 Hz, 1H, H-200), 3.17 (m, 2H, H-20,
H-300), 3.13 (dd, J ¼ 11.9, 4.6 Hz, 1H, H-3), 2.86 (dd, J ¼ 14.2,
3.7 Hz, 1H, H-18), 2.03 (dt, J ¼ 13.7, 3.7 Hz, 1H, H-16), 1.15, 1.04,
0.95, 0.93, 0.91, 0.84, 0.80 (s each, 3 H each, CH3 ꢂ 7); 13C NMR
(CDCl3, 100 MHz): d 178.3 (C-28), 144.9 (C-13), 123.9 (C-12),
106.3 (C-10), 95.9 (C-100), 90.7 (C-3), 80.7, 78.5, 77.3, 76.4, 75.1,
73.6, 72.3, 71.0, 62.3, 57.1, 48.1, 47.5, 43.1, 42.6, 40.9, 40.2,
39.9, 37.7, 34.9, 34.0, 33.3, 33.1, 31.6, 28.9, 28.7, 27.1, 26.3,
24.3, 24.0, 19.4, 17.3, 17.0, 16.3; HRESIMS: m/z calcd for
1.24, 1.07, 0.99, 0.89, 0.87, 0.83 (s each, 3 H each, CH3 ꢂ 7). 13
C
NMR (100 MHz, C5D5N) d 176.3 (C-28), 144.1 (C-13), 122.5 (C-
12), 104.7 (C-100), 102.9 (C-1000), 95.7 (C-10), 89.7 (C-3), 78.6, 78.3,
77.2, 76.9, 75.5, 74.3, 73.5, 72.5, 71.3, 71.0, 70.6, 69.1, 67.3,
62.0, 56.0, 49.7, 48.3, 47.1, 46.3, 42.2, 41.8, 40.0, 39.7, 38.9,
37.3, 34.1, 33.2, 32.7, 30.7, 28.4, 28.5, 26.8, 26.1, 23.9, 23.8,
23.4, 18.7, 17.6, 17.0, 15.9; HRESIMS: m/z calcd for
C
41H66O12Na [MþNaþ] 773.4446; found: 773.4426.
C
48H78O17Na [MþNaþ] 949.5131; found: 949.5150.
Oleanolic acid, 28-O-b-D-glucopyranosyl ester (6)
Yield: 95%; 1H NMR (CD3OD, 400 MHz): d 5.43 (d, J ¼ 8.0 Hz,
1H, H-100), 5.23 (t, J ¼ 3.7 Hz, 1H, H-12), 3.85 (t, J ¼ 9.9 Hz, 1 H,
H-40), 3.65 (dd, J ¼ 12.3, 3.4 Hz, 1 H, H-600-1), 3.38–3.42 (m, 2 H,
H-500, H-600-2), 3.31 (t, J ¼ 9.9, 8.0 Hz, 1H, H-200), 3.21 (t,
J ¼ 9.9 Hz, 1H, H-300), 3.17 (dd, J ¼ 11.9, 3.7 Hz, 1H, H-3), 2.83
(dd, J ¼ 14.7, 3.7 Hz, 1H, H-18), 1.13, 1.01, 0.95, 0.93, 0.89, 0.84,
0.79, (s each, 3 H each, CH3 ꢂ 7); 13C NMR (CDCl3, 100 MHz): d
178.3 (C-28), 144.3 (C-13), 123.7 (C-12), 95.9 (C-10), 90.6 (C-3),
78.0, 76.9, 75.1, 72.3, 62.3, 57.3, 48.3, 47.5, 43.1, 42.6, 40.7,
40.3, 39.9, 37.7, 35.1, 34.0, 33.3, 33.0, 31.6, 28.9, 28.6, 27.3,
26.3, 24.3, 24.0, 19.4, 17.3, 17.0, 16.1; HRESIMS: m/z calcd for
3-O-b-D-Glucopyranoside oleanolic acid (9)
23
Yield: 93%; [a]D ¼ þ 10.3 (c 0.85, CH3OH); 1H NMR (600 MHz,
DMSO-d6): d 12.05 (s, 1H, COOH), 5.16 (br s, 1H, H-12), 4.89,
0
0
0
4.86, 4.81 (brs each, 3 H, C2 -OH, C3 -OH, C4 -OH), 4.41 (br s, 1H,
0
0
C6 -OH), 4.12 (d, J ¼ 7.8 Hz, 1H, H-1 ), 3.67 (dd, J ¼ 11.7, 4.5 Hz,
1H, H-60-1), 3.45 (d, J ¼ 11.7, 6.1 Hz, 1H, H-60-2), 3.10 (dd,
J ¼ 11.7, 4.3 Hz, 1H, H-3), 3.02–3.07 (m, 3H, H-30, H-40, H-50),
3.00 (m, 1H, H-20), 2.78 (dd, J ¼ 13.7, 3.7 Hz, 1H, H-18), 1.08,
0.97, 0.88, 0.86, 0.85, 0.75, 0.66 (s each, 3 H each, CH3 ꢂ 7); 13
C
NMR (100 MHz, CD3OD): d 180.3 (C-28), 145.3 (C-13), 123.7 (C-
12), 106.7 (C-10), 90.9 (C-3), 78.5, 77.7, 75.6, 71.3, 62.8, 56.9,
52.0, 47.7, 47.5, 42.9, 42.7, 40.9, 40.6, 40.3, 39.8, 37.7, 35.1,
34.9, 34.3, 33.9, 33.6, 31.6, 30.5, 28.5, 27.7, 27.3, 26.5, 25.7,
24.6, 24.3, 19.3, 17.9, 16.1, 15.3; HRESIMS: m/z calcd for
C
36H58O8Na [MþNaþ] 641.4024; found: 641.4037.
Oleanolic acid, 28-O-b-D-glucopyranosyl-(1!6)-b-D-
glucopyranosyl ester (7)
C
36H58O8Na [MþNa]þ 641.4024; found: 641.4011.
Yield: 89%; 1H NMR (400 MHz, C5D5N) d: 6.21 (d, J ¼ 8.0 Hz, 1H,
H-100), 5.36 (t, J ¼ 3.7 Hz, 1H, H-12), 5.01 (d, J ¼ 7.9 Hz, 1H, H-10),
4.69 (t, J ¼ 9.7 Hz, 1H, H-40), 4.43 (t, J ¼ 9.7 Hz, 1H, H-300), 4.33
(dd, J ¼ 12.1, 3.3 Hz, 1H, H-60-1), 4.27 (t, J ¼ 9.7 Hz, 1H, H-30),
4.23 (dd, J ¼ 9.7, 8.0 Hz, 1H, H-200), 4.21 (t, J ¼ 9.7 Hz, 1H, H-400),
4.15 (dd, J ¼ 12.1, 5.1 Hz, 1H, H-60-2), 4.13 (m, 1H, H-50), 4.10
(dd, J ¼ 12.3, 3.3 Hz, 1H, H-600-1), 4.05 (m, 1 H, H-500), 3.97 (dd,
J ¼ 9.7, 7.9 Hz, 1H, H-20), 3.80 (dd, J ¼ 12.3, 5.0 Hz, 1H, H-600-2),
3.35 (dd, J ¼ 11.9, 3.7 Hz, 1H, H-3), 3.16 (dd, J ¼ 13.7, 4.1 Hz, 1H,
H-18), 1.26, 1.24, 1.05, 0.99, 0.90, 0.87, 0.86 (s each, 3 H each,
CH3 ꢂ 7); 13C NMR (100 MHz, C5D5N) d 176.1 (C-28), 144.5 (C-
13), 122.3 (C-12), 104.5 (C-100), 95.9 (C-10), 89.9 (C-3), 78.0, 76.9,
75.6, 74.1, 73.5, 72.5, 71.1, 70.6, 69.1, 62.0, 56.0, 49.7, 48.6,
47.1, 46.3, 42.2, 41.8, 40.0, 39.7, 38.9, 37.5, 34.1, 33.2, 32.7,
30.7, 28.9, 28.5, 26.5, 26.3, 23.7, 23.8, 23.4, 18.7, 17.6, 17.0,
3-O-b-D-Xylopyranoside oleanolic acid (10)
23
Yield: 93%; [a]D ¼ þ 23.6 (c 1.01, CH3OH); 1H NMR (600 MHz,
DMSO-d6): d 12.03 (s, 1H, COOH), 5.19 (br s, 1H, H-12), 4.90,
0
0
0
4.87, 4.85 (brs each, 3 H, C2 -OH, C3 -OH, C4 -OH), 4.23 (d,
J ¼ 7.6 Hz, 1H, H-10), 3.53 (dd, J ¼ 12.5, 3.1 Hz, 1H, H-50-1), 3.31
(dd, J ¼ 12.5, 5.1 Hz, 1H, H-50-2), 3.09 (dd, J ¼ 11.7, 4.3 Hz, 1H,
H-3), 2.95–3.06 (m, 3H, H-20, H-30, H-40), 2.76 (dd, J ¼ 13.7,
3.7 Hz, 1H, H-18), 1.09, 0.97, 0.91, 0.87, 0.76, 0.75, 0.65 (s each,
3H each, CH3 ꢂ 7); 13C NMR (100 MHz, CD3OD): d 180.1 (C-28),
145.3 (C-13), 123.5 (C-12), 107.5 (C-10), 89.9 (C-3), 78.5, 75.6,
71.3, 67.3, 55.9, 48.1, 46.7, 46.5, 42.3, 42.0, 40.1, 40.0, 39.9,
39.8, 37.7, 35.1, 34.9, 34.3, 33.9, 33.6, 31.6, 30.7, 28.3, 27.7,
27.3, 26.5, 25.9, 24.6, 23.9, 18.3, 17.5, 17.1, 15.6; HRESIMS: m/z
calcd for C35H56O7Na [MþNa]þ 611.3918; found: 611.3931.
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Archiv der Pharmazie
3-O-b-D-Galactopyranoside oleanolic acid (11)
0.83, 0.83, 0.72, 0.69 (s each, 3H each, CH3 ꢂ 7); 13C NMR
(100 MHz, CD3OD): d 180.1 (C-28), 145.0 (C-13), 123.7 (C-12),
102.3 (C-10), 86.5 (C-3), 75.3, 73.1, 72.4, 71.5, 59.3, 53.1, 47.8,
46.5, 44.9, 43.6, 41.7, 40.3, 39.7, 39.1, 37.9, 35.3, 34.7, 34.1,
33.9, 33.3, 31.6, 30.1, 28.9, 28.7, 27.9, 26.7, 26.3, 26.1, 25.7,
24.3, 24.0, 22.5, 19.5, 18.7, 17.2, 16.8, 15.3; HRESIMS: m/z calcd
for C36H59O7 [MþH]þ 603.4253; found: 603.4242.
23
Yield: 87%; [a]D ¼ þ 20.3 (c 0.76, CH3OH); 1H NMR (600 MHz,
DMSO-d6): d 12.06 (s, 1H, COOH), 5.17 (br s, 1H, H-12), 4.80,
0
0
0
0
4.76, 4.64, 4.40 (brs each, 4 H, C2 -OH, C3 -OH, C4 -OH, C6 -OH),
4.13 (d, J ¼ 6.9 Hz, 1H, H-10), 3.63 (dd, J ¼ 12.1, 3.5 Hz, 1H, H-60-
1), 3.52 (dd, J ¼ 12.1, 5.7 Hz, 1H, H-60-2), 3.44 (m, 1H, H-50),
3.21–3.26 (m, 3H, H-20, H-30, H-40), 3.11 (dd, J ¼ 11.7, 4.1 Hz, 1H,
H-3), 2.73 (dd, J ¼ 13.7, 3.3 Hz, 1H, H-18), 1.09, 0.98, 0.90, 0.87,
0.76, 0.73, 0.69 (s each, 3H each, CH3 ꢂ 7); 13C NMR (100 MHz,
CD3OD): d 180.0 (C-28), 144.1 (C-13), 123.1 (C-12), 106.1 (C-10),
89.7 (C-3), 75.3, 73.6, 71.3, 68.3, 60.6, 55.1, 47.1, 45.7, 46.5,
42.3, 41.6, 40.1, 39.9, 39.3, 38.7, 38.3, 36.1, 34.9, 33.3, 32.9,
32.6, 31.6, 30.5, 28.3, 27.7, 27.1, 26.5, 25.7, 24.6, 23.9, 22.6,
17.9, 16.7, 15.3; HRESIMS: m/z calcd for C36H58O8Na [MþNa]þ
641.4023; found: 641.4037.
3-O-b-D-Glucopyranosyl-(1!4)-b-D-glucopyranoside
oleanolic acid (15)
25
Yield: 90%; [a]D ¼ þ 81.67 (c 1.20, CHCl3); 1H NMR (600 MHz,
DMSO-d6): d 12.02 (s, 1H, COOH), 5.19 (br s, 1H, H-12), 5.13 (d,
00
0
J ¼ 4.9 Hz, 1H, C2 -OH), 5.03 (d, J ¼ 5.1 Hz, 1H, C2 -OH), 4.83 (d,
0
00
00
00
J ¼ 5.3 Hz, 1H, C3 -OH), 4.69–4.73 (m, 3H, C3 -OH, C4 -OH, C6
-
0
OH), 4.51 (t, J ¼ 6.0 Hz, 1H, C6 -OH), 4.23 (d, J ¼ 7.7 Hz, 1H, H-
10), 4.19 (d, J ¼ 7.5 Hz, 1H, H-100), 3.53–3.63 (m, 6 H, H-300, H-400,
H-600-1, H-600-2, H-60-2, H-60-1), 3.35–3.40 (m, 2 H, H-30, H-40),
3.23–3.31 (m, 2 H, H-50, H-500), 3.07 (dd, J ¼ 12.1, 3.7 Hz, 1H, H-
3), 2.95 (m, 1 H, H-20), 2.76 (dd, J ¼ 13.7, 4.3 Hz, 1H, H-18), 1.03,
3-O-a-D-Mannopyranoside oleanolic acid (12)
23
1
Yield: 88%; [a]D ¼ þ 33.1 (c 1.00, CHCl3); H NMR (600 MHz,
DMSO-d6): d 12.05 (s, 1H, COOH), 5.21 (t, J ¼ 3.2Hz, 1H, H-12),
4.79 (s, 1H, H-1 ), 4.73 (d, J ¼ 4.8 Hz, 1H, C3 -OH), 4.63 (d,
0.99, 0.88, 0.85, 0.83, 0.75, 0.69 (s each, 3H each, CH3 ꢂ 7); 13
C
0
0
0
0
J ¼ 4.5 Hz, 1H, C2 -OH), 4.51 (d, J ¼ 5.1 Hz, 1H, C4 -OH), 4.33 (t,
NMR (100 MHz, CD3OD): d 179.1 (C-28), 143.9 (C-13), 122.9 (C-
12), 105.1 (C-10), 104.1 (C-100), 89.9 (C-3), 79.3, 76.0, 75.1, 74.6,
73.9, 73.3, 71.6, 68.7, 63.7, 60.3, 56.3, 50.1, 46.5, 46.2, 41.7,
41.5, 39.4, 38.9, 38.5, 36.7, 33.8, 33.5, 32.6, 32.3, 31.5, 28.9,
27.2, 26.4, 25.7, 25.1, 24.4, 23.3, 22.7, 22.3, 18.3, 16.7, 16.1,
15.0, 14.9; HRESIMS: m/z calcd for C42H68O13Na [MþNa]þ
803.4551; found: 803.4567.
0
0
0
J ¼ 5.5 Hz, 1H, C5 -OH), 3.60 (m, 1H, H-6 -1), 3.50 (m, 1H, H-2 ),
3.31–3.37 (m, 4H, H-30, H-40, H-50), 3.13 (dd, J ¼ 11.7, 3.7 Hz, 1H,
H-3), 2.76 (dd, J ¼ 13.7, 4.3 Hz, 1H, H-18), 1.09, 0.93, 0.89, 0.87,
0.85, 0.73, 0.65 (s each, 3 H each, CH3 ꢂ 7); 13C NMR (100 MHz,
CD3OD): d 180.3 (C-28), 145.1 (C-13), 123.5 (C-12), 97.6 (C-10),
83.1 (C-3), 75.1, 73.1, 72.7, 68.9, 62.5, 56.9, 47.9, 43.6, 41.7,
40.6, 40.1, 39.4, 39.0, 38.1, 35.1, 34.7, 34.5, 33.9, 32.6, 31.5,
30.1, 29.5, 28.53, 27.8, 26.5, 25.7, 24.5, 23.9, 21.6, 19.7, 18.1,
17.3, 16.1; HRESIMS: m/z calcd for C36H58O8Na [MþNa]þ
641.4024; found: 641.4029.
3-O-b-D-Galactopyranosyl-(1!4)-b-D-glucopyranoside
oleanolic acid (16)
23
1
Yield: 93%; [a]D ¼ þ 45.7 (c 1.01, CHCl3); H NMR (600 MHz,
DMSO-d6): d 12.01 (s, 1H, COOH), 5.17 (t, J ¼ 3.7 Hz, 1H, H-12),
0
0
00
00
3-O-a-L-Rhamnopyranoside oleanolic acid (13)
4.73–4.87 (m, 4H, C2 -OH, C3 -OH, C2 -OH, C3 -OH), 4.65 (d,
25
Yield: 89%; [a]D ¼ 13.5 (c 1.15, CHCl3); 1H NMR (600 MHz,
J ¼ 5.5 Hz, 1H, C4 -OH), 4.55 (d, J ¼ 5.1 Hz, 1H, C4 -OH), 4.31 (d,
0
00
00
0
DMSO-d6): d 12.01 (s, 1H, COOH), 5.15 (t, J ¼ 3.6 Hz, 1H, H-12),
J ¼ 4.7 Hz, 1H, C6 -OH), 4.27 (d, J ¼ 4.9 Hz, 1H, C6 -OH), 4.24 (d,
J ¼ 7.6 Hz, 1H, H-10), 4.21 (d, J ¼ 7.5 Hz, 1H, H-100), 3.72 (dd,
J ¼ 12.1, 3.9 Hz, 1H, H-60-1), 3.63 (dd, J ¼ 12.3, 4.5 Hz, 1H, H-600-
1), 3.57–3.61 (m, 2H, H-50, H-500), 3.49 (dd, J ¼ 12.3, 6.0 Hz, 1H,
H-600-2), 3.43 (dd, J ¼ 12.1, 5.3 Hz, 1H, H-60-2), 3.21–3.25 (m, 3H,
30, 300, 400), 3.09 (dd, J ¼ 11.7, 3.7 Hz, 1H, H-3), 3.03–3.07 (m, 3H,
20, 200, 40), 3.38 (t, J ¼ 3.6 Hz, 1H, H-40), 3.23 (m, 1H, H-30), 3.17
(m, 1H, H-20), 2.78 (dd, J ¼ 12.1, 3.7 Hz, 1H, H-18), 1.03, 0.89,
0.86, 0.85, 0.79, 0.72, 0.63 (s each, 3 H each, CH3 ꢂ 7); 13C NMR
(100 MHz, CD3OD): d 179.8 (C-28), 144.7 (C-13), 122.9 (C-12),
105.2 (C-10), 103.9 (C-100), 88.3 (C-3), 81.5, 75.7, 75.3, 74.9, 73.6,
72.9, 71.0, 68.9, 61.1, 56.3, 51.1, 47.3, 46.7, 45.9, 41.6, 40.3,
39.5, 38.1, 37.9, 35.7, 34.2, 34.1, 33.7, 33.1, 31.6, 30.1, 28.9,
28.7, 27.9, 26.7, 26.3, 26.1, 25.7, 24.3, 24.0, 23.1, 22.5, 19.5,
18.3, 17.1, 16.8, 15.3; HRESIMS: m/z calcd for C42H68O13Na
[MþNa]þ 803.4553; found: 803.4539.
0
0
4.73 (br s, 1H, H-1 ), 4.69 (d, J ¼ 4.9 Hz, 1H, C2 -OH), 4.59 (d,
0
0
J ¼ 5.0 Hz, 1H, C3 -OH), 4.50 (d, J ¼ 5.6 Hz, 1H, C4 -OH), 3.42–
3.56 (m, 3H, H-20, H-30, H-40), 3.19 (m, 1H, H-50), 3.07 (dd,
J ¼ 12.3, 4.3 Hz, 1H, H-3), 2.77 (dd, J ¼ 13.7, 3.7 Hz, 1H, H-18),
1.13 (d, J ¼ 5.9 Hz, 3 H, H-60), 1.08, 0.90, 0.89, 0.87, 0.71, 0.70,
0.69 (s each, 3H each, CH3 ꢂ 7); 13C NMR (100 MHz, CD3OD): d
180.1 (C-28), 144.7 (C-13), 122.9 (C-12), 102.9 (C-10), 87.1 (C-3),
72.3, 71.9, 70.9, 68.9, 62.5, 54.9, 47.3, 45.9, 43.6, 41.3, 40.9,
40.1, 39.4, 38.8, 38.5, 37.9, 36.4, 35.1, 34.5, 33.5, 32.3, 31.7,
30.5, 28.5, 27.8, 26.5, 25.9, 24.5, 23.9, 23.0, 22.3, 19.7, 18.1,
17.8, 17.1, 16.1; HRESIMS: m/z calcd for C36H59O7 [MþH]þ
603.4255; found: 603.4261.
3-O-a-L-Fucopyranoside oleanolic acid (14)
25
Yield: 87%; [a]D ¼ 30.9 (c 1.15, CH3OH); 1H NMR (600 MHz,
DMSO-d6): d 12.03 (s, 1H, COOH), 5.20 (t, J ¼ 3.6 Hz, 1H, H-12),
0
0
Biological evaluations
Assay for a-glucosidase inhibitory activities
Inhibitory a-glucosidase activities were determined spectro-
photometrically in a 96-well microtiter plate based on p-
nitrophenyl-a-D-glucopyranoside (pNPG) as a substrate. In
4.61 (d, J ¼ 5.0 Hz, 1H, C2 -OH), 4.50 (d, J ¼ 4.9 Hz, 1H, C3 -OH),
0
0
4.31 (d, J ¼ 4.7 Hz, 1H, C4 -OH), 4.09 (d, J ¼ 7.5 Hz, 1H, H-1 ),
3.40 (dd, J ¼ 12.1, 5.6 Hz, 1H, H-50), 3.38 (t, J ¼ 3.6 Hz, 1H, H-40),
3.23 (m, 1H, H-30), 3.17 (m, 1H, H-20), 3.06 (dd, J ¼ 11.9, 4.3 Hz,
1H, H-3), 2.78 (dd, J ¼ 12.1, 3.7 Hz, 1H, H-18), 0.99, 0.89, 0.86,
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Archiv der Pharmazie
brief, 20 mL of enzyme solution [0.8 U/mL a-glucosidase in
0.01 M potassium phosphate buffer (pH 6.8) containing 0.2%
of BSA] and 120 mL of the synthetic compound in 0.5% DMSO
of 0.01 M potassium phosphate buffer were mixed, and was
preincubated at 37°C prior to initiation of the reaction by
adding the substrate. After 15 min of preincubation, pNPG
solution (20 mL) [5.0 mM pNPG in 0.1 M potassium phosphate
buffer (pH 6.8)] was added and then incubated together at
37°C. After 15 min of incubation, 0.2 M Na2CO3 (80 mL) in 0.1 M
potassium phosphate buffer was added to the test tube to
stop the reaction. Acarbose was used as a positive control. The
increment in absorption at 410 nm due to the hydrolysis of
pNPG by a-glucosidase was monitored continuously with an
auto multi-functional microplate reader (BIORAD680).
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Assay for a-amylase inhibitory activities
The a-amylase inhibitory activities were measured with the
method reported by Xiao et al. [47] and Yoshikawa et al. [48]
with slight modifications. Substrate was prepared by heating
starch (250 mg) in 12 mL of 0.4 M NaOH solution for 5 min at
100°C, and then cooled to 0°C and adjusted to pH 7 with 2 M
HCl. Sample solutions were prepared by dissolving each
solution in acetate buffer (pH 6.5). The sample (20 mL) and the
substrate (40 mL) were mixed in a microplate well. After
preincubation at 37°C for 15 min, 5 mg/mL a-amylase solution
(20 mL) was added and the solution was incubated at 37°C for
15 min. The reaction was stopped by adding 50 mL 1 M HCl,
and then 50 mL iodine solution was added. The absorbances
were measured at 650 nm by a microplate reader. Acarbose
was used as a positive control.
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This work was financially supported by China Postdoctoral
Science Foundation funded project (No. 2015M572596), Hong
Kong Scholars Program Foundation (XJ2014014), the Post-
doctoral Science Foundation of Northwest University (Nos.
334100018 and 334100030), the Foundation of Outstanding
Young Academic backbone Supporting Program of North-
west University (No. 338050029), the Foundation of Natural
Science Basic Research Plan of Shaanxi Province of China
(2014JQ2069), Shaanxi Youth Science and Technology Star
Project (2015KJXX-28) and the Scientific Research Foundation
of Northwest University (No. 14NW08).
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The authors have declared no conflicts of interest.
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