α -Amylase and α -Glucosidase Inhibitory Saponins from Polyscias fruticosa Leaves

Article abstract of DOI:10.1155/2016/2082946

Three bisdesmosidic saponins 3-O-[β-D-glucopyranosyl-(1 → 4)-β-D-glucuronopyranosyl] oleanolic acid 28-O-β-D-glucopyranosyl ester (1), polyscioside D (2), and 3-O- { β -D-glucopyranosyl-(1 → 2)-[β-D-glucopyranosyl-(1 → 4)]-β-D- g l u c u r o n o p y r a n o s y l } oleanolic acid 28-O-β-D-glucopyranosyl-(1 → 2)-β-D-galactopyranosyl ester (3) were isolated from a methanol extract of Polyscias fruticosa (L.) Harms leaves. Compound 1 was obtained as a main constituent and compound 3 was reported for the first time and named as polyscioside I. Saponin 1 inhibited porcine pancreas α-amylase and yeast α-glucosidase activities while 2 and 3 were inactive. Synergistic inhibitory effect on α-amylase was observed from the combination of low concentrations of 1 and acarbose. The findings suggest the use of P. fruticosa and its major saponin 1 for the prevention and treatment of diabetes and its complications.

Full text of DOI:10.1155/2016/2082946

Hindawi Publishing Corporation
Journal of Chemistry
Volume 2016, Article ID 2082946, 5 pages
http://dx.doi.org/10.1155/2016/2082946
Research Article
ꢀ-Amylase and ꢀ-Glucosidase Inhibitory Saponins from
Polyscias fruticosa Leaves
Tran Thi Hong Hanh, Nguyen Hai Dang, and Nguyen Tien Dat
Institute of Marine Biochemistry, Vietnam Academy of Science and Technology, 18-Hoang Quoc Viet, Cau Giay,
Hanoi 100000, Vietnam
Correspondence should be addressed to Nguyen Tien Dat; ngtiend@imbc.vast.vn
Received 21 February 2016; Accepted 17 April 2016
Academic Editor: Nick Kalogeropoulos
Copyright © 2016 Tran i Hong Hanh et al. is is an open access article distributed under the Creative Commons Attribution
License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly
cited.
ree bisdesmosidic saponins 3-O-[ꢀ-D-glucopyranosyl-(1→4)-ꢀ-D-glucuronopyranosyl] oleanolic acid 28-O-ꢀ-D-glucopyran-
osyl ester (1), polyscioside D (2), and 3-O-{ꢀ-D-glucopyranosyl-(1→2)-[ꢀ-D-glucopyranosyl-(1→4)]-ꢀ-D-glucuronopyranosyl}
oleanolic acid 28-O-ꢀ-D-glucopyranosyl-(1→2)-ꢀ-D-galactopyranosyl ester (3) were isolated from a methanol extract of Polyscias
fruticosa (L.) Harms leaves. Compound 1 was obtained as a main constituent and compound 3 was reported for the first time and
named as polyscioside I. Saponin 1 inhibited porcine pancreas ꢁ-amylase and yeast ꢁ-glucosidase activities while 2 and 3 were
inactive. Synergistic inhibitory effect on ꢁ-amylase was observed from the combination of low concentrations of 1 and acarbose.
e findings suggest the use of P. fruticosa and its major saponin 1 for the prevention and treatment of diabetes and its complications.
1. Introduction
Our search for antidiabetic agents of natural origin iden-
tified a methanol extract of P. fruticosa leaves that showed
significant inhibitory activity against ꢁ-glucosidase and
ꢁ-amylase. A phytochemical investigation of the methanol
extract of P. fruticosa leaves led to the isolation of three
bisdesmosidic saponins (Figure 1). eir structures were
elucidated as 3-O-[ꢀ-D-glucopyranosyl-(1→4)-ꢀ-D-glucu-
ronopyranosyl] oleanolic acid 28-O-ꢀ-D-glucopyranosyl
ester (1) [6, 8], 3-O-{ꢀ-D-glucopyranosyl-(1→2)-[ꢀ-D-gluco-
pyranosyl-(1→4)]-ꢀ-D-glucuronopyranosyl} oleanolic acid
28-O-ꢀ-D-glucopyranosyl ester or polyscioside D (2) [6],
and 3-O-{ꢀ-D-glucopyranosyl-(1→2)-[ꢀ-D-glucopyranosyl-
(1→4)]-ꢀ-D-glucuronopyranosyl} oleanolic acid 28-O-ꢀ-D-
glucopyranosyl-(1→2)-ꢀ-D-galactopyranosyl ester (3), new-
ly named polyscioside I. All compounds were evaluated for
their inhibitory activity against porcine pancreas ꢁ-amylase
and yeast ꢁ-glucosidase.
Polyscias fruticosa (L.) Harms (synonyms Nothopanax fru-
ticosus (L.) Miq. and Panax fruticosus L.), belonging to the
family Araliaceae, is widely used in Vietnam as a tonic
agent for the treatment of ischemia and inflammation and
to increase blood flow in the brain. e leaves of this
plant are also eaten as a salad [1]. Previous studies showed
that P. fruticosa leaf extracts exhibited antipyretic, anti-
inflammatory, analgesic, and molluscicidal properties [2, 3]
and antidiabetic activity [4]. A few reports on the chemical
composition of this plant indicated the presence of polyacety-
lene [5] and saponins [6]. Diabetes is a group of metabolic
diseases characterized by chronic hyperglycemia resulting
from deficiency in insulin secretion or action. Recent reports
revealed that a high postprandial plasma glucose level is more
harmful than fasting blood glucose. erefore, it is important
to control the postprandial blood glucose level to reduce
complications and mortality. One therapeutic approach for
treating diabetes is to decrease postprandial glycemia by
inhibiting enzymes responsible for carbohydrate hydrolysis,
such as ꢁ-glucosidase and ꢁ-amylase [7].
2. Experimental
2.1. General Experimental Procedures. Optical rotation values
were recorded a JASCO P-2000 digital polarimeter (JASCO,

2
Journal of Chemistry
30
19
29
6ꢀ
4ꢀ
O
HO
6ꢀꢀ
20
HO
HO
6ꢀ
4ꢀ
O
21
22
HO
6ꢀꢀ
O
O
1ꢀꢀ
HO
HO
O
O
O
O
1ꢀ
1ꢀꢀ
HO
HO
3ꢀ
18
O
HO
1ꢀ
12
OH
3ꢀꢀ
O
28
11
HO
13
14
17
16
3ꢀ
25
24
OH
26
OH
3ꢀꢀ
HO
15
6ꢀꢀꢀꢀ
O
1
9
1ꢀꢀꢀꢀ
S1
OR2
HO
2
3
10
5
8
7
S2
OH
HO
OH
3ꢀꢀꢀꢀ
27
4
6
R1O
6ꢀ
3ꢀ
HO
OH
4ꢀ
23
6ꢀꢀꢀ
O
4ꢀꢀꢀ
HO
O
ꢀ
HO
OH1
1: R1 = S1, R2 = S3
2: R1 = S2, R2 = S3
3: R1 = S2, R2 = S4
1ꢀꢀꢀ
HO
6ꢀꢀꢀꢀꢀ
3ꢀꢀꢀ
O
HO
HO
S3
O
1ꢀꢀꢀꢀꢀ
HO
OH S4
3ꢀꢀꢀꢀꢀ
30
29
20
19
21
22
18
15
12
26
O
28
11
13
14
17
16
25
1
9
10
2
8
7
O
O
6ꢀ
27
HO
3
5
6ꢀꢀ
HO
HO
4ꢀ
4
6
O
O
O
OH
6ꢀꢀꢀ
1ꢀꢀ
O
OH
HO
23
24
3ꢀ
HO
ꢀ
O 1
OH
HO
3ꢀꢀ
O
4ꢀꢀꢀ
HO
6ꢀꢀꢀꢀꢀ
1ꢀꢀꢀ
6ꢀꢀꢀ
3ꢀꢀꢀ
O
O
1ꢀꢀꢀꢀ
HO
HO
HO
O
HO
OH
1ꢀꢀꢀꢀꢀ
3ꢀꢀꢀꢀ
HO
OH
3
Figure 1: Structure of compounds 1–3 and key HMBC correlations of 3.
Tokyo, Japan). NMR experiments were carried out on a
Bruker AM500 FT-NMR spectrometer (Bruker, Rheinstet-
ten, Germany) using tetramethylsilane (TMS) as internal
standard. e HR-ESI-MS was recorded on API Q-STAR
PULSAR I of Applied Biosystem. e bioassay absorbance
was read by xMark Microplate Spectrophotometer reader
(Bio-Rad Laboratories, USA).
and 92 g of hexane and ethyl acetate residues, respectively.
e water residue was filtered through a Diaion HP-20
column eluted by 0%, 25%, and 100% methanol in water.
e 100% methanol-eluted fraction was diluted with water
(1 : 1 v/v) and kept overnight at 4∘C. Compound 1 as white
crystals (500 mg) was obtained by filtering. e filtrate was
concentrated and chromatographed on a silica gel column
eluted with a gradient of 0%, 50%, and 100% methanol in
ethyl acetate to afford three fractions, F1–F3. Fraction F2 was
fractionated on a silica gel column eluted with chloroform-
methanol-water (50 : 10 : 1 v/v) to afford compound 2 (10 mg).
Compound 3 (20.5 mg) was purified from F3 by a RP-18
column (methanol-water 1 : 1 v/v).
2.2. Plant Material. e leaves of Polyscias fruticosa were
collected in Tien Hai, ai Binh, in July 2011 and identified by
Professor Tran Huy ai, Institute of Ecology and Biological
Resources, Vietnam Academy of Science and Technology.
e voucher specimens were deposited at the herbarium of
the Institute of Ecology and Biological Resources.
24
Polyscioside I (3). White powder, mp. >300∘C. [ꢁꢂ
= +12.4
(c 0.1, MeOH). 1H NMR (500 MHz, DMSO-d ):Dꢃ 0.67 (3H,
6
2.3. Extraction and Isolation. e air-dried and powdered P.
br s, H-26), 0.73 (3H, br s, H-24), 0.85 (6H, br s, H-25, 29),
fruticosa leaves (2.0 kg) were extracted with methanol (5 L ×
0.87 (3H, br s, H-30), 0.97 (3H, br s, H-23), 1.06 (3H, br s,
3 times) at room temperature. e combined extracts were
H-27), 3.01 (1H, m, H-3), 3.60 (1H, m, H-2ꢀꢀꢀ), 4.25 (1H, ꢄ,
concentrated to give 300 g of crude extract, which was then
J = 7.0 Hz, H-1ꢀꢀ), 4.29 (1H, ꢄ, J = 6.5 Hz, H-1ꢀ), 4.62 (1H, ꢄ,
resuspended in water (1.5 L) and successively partitioned with
J = 7.5 Hz, H-1ꢀꢀꢀꢀꢀ), 4.48 (1H, ꢄ, J = 7.0 Hz, H-1ꢀꢀꢀꢀ), 5.14 (1H,
m, H-11), 5.30 (1H, ꢄ, J = 8.0 Hz, H-1ꢀꢀꢀ). 13C NMR (125 MHz,
hexane and ethyl acetate (each 0.5 L × 3 times) to obtain 75

Journal of Chemistry
3
Table 1: 13C-NMR data of compounds 1–3 measured in DMSO-ꢄ6.
DMSO-d ): see Table 1. HR-ESI-MS (positive): ꢅꢆ/ 1281.6127
6
[M + H]+, (calcd. 1281.6116 for C H O ).
60 97 29
C
1
1
2
38.1
3
38.1
C
1
2
3
38.1
3-O-GluA
Acid Hydrolysis of 3. Compound 3 (10 mg) was heated in
2 N HCl (5 mL) at 80∘C for 2 h, and then the solution was
extracted with ethyl acetate (2 mL × 3). e sugar products
in the aqueous layer were identified as glucose (Rf 0.65),
galactose (Rf 0.60), and glucuronic acid (Rf 0.12) in compar-
ison with standards by silica gel thin layer chromatography
(developed with acetone-methanol-water 10 : 9 : 1 and sprayed
by 10% sulfuric acid solution containing 2% vanillin). Afer
preparative thin layer chromatography, the optical rotation of
each sugar fraction was checked. e positive optical rotation
values led to the assignment of D-glucuronic acid, D-glucose,
and D-galactose [9].
2
3
4
5
6
7
8
9
25.4
87.9
38.7
55.0
17. 7
22.9
88.4
38.6
55.0
17. 7
25.4
88.3
38.6
55.0
17. 7
1ꢀ
2ꢀ
105.0 103.6 103.6
73.5
75.4
82.9
75.3
79.3
75.4
82.3
75.1
79.5
75.3
82.3
75.1
3ꢀ
4ꢀ
5ꢀ
6ꢀ
32.2
32.3
32.7
172.0 171.8 171.5
40.0 40.7 40.0
4ꢀ-O-Glu
1ꢀꢀ
47. 0
47. 0
36.2
22.9
47. 0
36.2
22.9
103.8 103.3 103.3
10 36.2
11 22.9
2ꢀꢀ
73.4
76.3
70.0
77.0
61.1
73.5
76.2
69.9
77.0
61.1
73.6
76.3
69.9
77.0
61.0
3ꢀꢀ
2.4. Assay for ꢁ-Glucosidase Inhibition. e yeast ꢁ-glu-
12 121.6 121.7 121.5
13 143.4 143.4 143.5
4ꢀꢀ
cosidase (G0660, Sigma-Aldrich) enzyme inhibition assay
5ꢀꢀ
was performed by the digestion of p-nitrophenyl-ꢁ-D-
glucopyranoside [10]. e sample solution (2 ꢇL dissolved in
DMSO) and 0.5 U/mL ꢁ-glucosidase (40 ꢇL) were mixed in
120 ꢇL of 0.1 M phosphate buffer (pH 7.0). Afer 5 min prein-
cubation, 5 mM p-nitrophenyl-ꢁ-D-glucopyranoside solu-
tion (40 ꢇL) was added and the solution was incubated at
37∘C for 30 min. e absorbance of released 4-nitrophenol
was measured at 405 nm by using a microplate reader (Molec-
ular Devices, CA). Acarbose was used as positive control
14 41.2
15 27.1
16 22.5
17 45.9
41.2
27.7
22.5
45.9
41.1
27.9
21.9
45.8
6ꢀꢀ
2ꢀ-O-Glu
1ꢀꢀꢀꢀ
2ꢀꢀꢀꢀ
3ꢀꢀꢀꢀ
4ꢀꢀꢀꢀ
5ꢀꢀꢀꢀ
6ꢀꢀꢀꢀ
—
—
—
—
—
—
103.1 103.2
73.5
76.1
70.1
76.8
60.9
73.6
76.0
70.0
76.7
61.0
18 40.7 40.7 40.7
19 45.5
20 30.2
21 33.2
22 31.6
23 27.6
24 16.6
25 15.1
26 16.4
27 25.4
45.5
30.3
33.2
31.6
27.7
16.1
15.1
45.5
30.2
33.2
(IC 650.4 ꢇg/mL).
50
32.3 28-O-Glc/Gal
2.5. Assay for ꢁ-Amylase Inhibition. e porcine pancreas ꢁ-
amylase (A3176, Sigma-Aldrich) enzyme inhibitory activity
was measured using the method reported by Ashok Kumar
et al. [11] with slight modifications. Substrate was pre-
pared by dissolving 2-chloro-4-nitrophenyl-ꢁ-D-maltotrio
side (93834, Sigma-Aldrich) in phosphate buffer (pH 7.0). e
sample (2 ꢇL dissolved in DMSO) and 0.5 unit/mL ꢁ-amylase
(50 ꢇL) were mixed in 100 ꢇL of 0.1 M phosphate buffer (pH
7.0). Afer 5 min preincubation, substrate solution (50 ꢇL) was
added and the solution was incubated at 37∘C for 15 min. e
absorbances were measured at 405 nm. Acarbose was used as
27.7
16.7
15.1
1ꢀꢀꢀ
2ꢀꢀꢀ
94.1
72.3
76.6
69.5
77.6
60.7
94.1
72.3
76.6
69.5
77.7
60.7
91.9
76.9
74.1
70.7
76.8
61.8
3ꢀꢀꢀ
16.9
25.5
16.7
25.5
4ꢀꢀꢀ
5ꢀꢀꢀ
28 175.2 175.2 175.2
6ꢀꢀꢀ
29 32.7
30 23.3
32.7
23.3
32.7
23.4
2ꢀꢀꢀ-O-Glu
1ꢀꢀꢀꢀꢀ
—
—
—
—
—
—
—
—
—
—
—
—
102.3
75.0
77.0
69.4
77.5
positive control (IC 57.6 ꢇg/mL).
2ꢀꢀꢀꢀꢀ
3ꢀꢀꢀꢀꢀ
4ꢀꢀꢀꢀꢀ
5ꢀꢀꢀꢀꢀ
6ꢀꢀꢀꢀꢀ
50
3. Results and Discussion
Compound 3 was obtained as a white crystal and its
high-resolution electrospray ionization mass spectrometry
spectrum revealed an ion peak at ꢅꢆ/ 1281.6127 [M + H]+,
60.5
corresponding to the molecular formula C H O . e
60 96 29
1H-NMR spectrum of 1 showed seven methyl groups at ꢃ
H
0.67 (3H, br s, H-26), 0.73 (3H, br s, H-24), 0.85 (6H, br
s, H-25, 29), 0.87 (3H, br s, H-30), 0.97 (3H, br s, H-23),
and 1.06 (3H, br s, H-27) and five glycosidic anomeric
seven CH , ten CH , five CH, and 8 C signals (Table 1).
3
2
e aglycone was identified as oleanolic acid, a common
triterpene of saponins in the genus Polyscias [6]. e ꢃ
C
protons at ꢃ 4.25 (1H, ꢄ, J = 7.0 Hz, H-1ꢀꢀ), 4.29 (1H, ꢄ, J =
values of C-3 at ꢃ 88.3 and C-28 at ꢃ 175.2 suggested that
H
C
compound 3 wasCa bisdesmosidic saponin with sugar units
6.5 Hz, H-1ꢀ), 4.48 (1H, ꢄ, J = 7.0 Hz, H-1ꢀꢀꢀꢀ), 5.30 (1H, ꢄ, J =
8.0 Hz, H-1ꢀꢀꢀ), and 4.62 (1H, ꢄ, J = 7.5 Hz, H-1ꢀꢀꢀꢀꢀ). e large
coupling constants of all anomeric protons indicated the
ꢀ-configuration of the sugars. 13C-NMR and DEPT spectra
indicated the presence of an olean triterpene skeleton with
attached to these positions. In addition, five sugar moieties
were recognized by five anomeric methine groups at ꢃ 103.6
C
(C-1ꢀ), 103.3 (C-1ꢀꢀ), 91.9 (C-1ꢀꢀꢀ), 103.2 (C-1ꢀꢀꢀꢀ), and 102.3
(C-1ꢀꢀꢀꢀꢀ). ese data are similar to those of compound 2,

4
Journal of Chemistry
Table 2: Inhibitory effects (IC
and ꢁ-glucosidase.
SD, ꢇg/mL) of 1–3 for ꢁ-amylase
except for the addition of a sugar moiety and the upfield shif
of the anomeric signal at ꢃ from 94.1 to 91.9. Acid hydrolysis
of compound 3 allowed tChe identification of D-glucuronic
acid, D-glucose, and D-galactose. Extensive analysis of the
HMBC spectrum of compound 3 allowed the position of
the sugar units to be determined. e couplings from H-1ꢀ
(ꢃ 4.29) to C-3 (ꢃ 88.3) indicated that the glucuronic and
50
Compounds
1
2
ꢁ-Amylase
ꢁ-Glucosidase
27.1 2.05
440.5 12.7
—
—
—
—
3
H
C
glucosyl moieties attached to C-3. e correlations from
Acarbose
57.6 4.12
650.4 33.4
H-1ꢀꢀ (ꢃ 4.25) to C-4ꢀ (ꢃ 82.3) and between H-4ꢀꢀꢀꢀ (ꢃ
H
C
H
4.48) and C-2ꢀ (ꢃ 79.5) confirmed that the two glucosyl
C
units were attached to C-2ꢀ and C-4ꢀ, respectively. Indeed, the
13C-NMR data of this fragment coincided well with the ꢀ-
D-glucopyranosyl-(1→2)-[ꢀ-D-glucopyranosyl-(1→4)]-ꢀ-D-
glucuronopyranosyl of compound 2 (Table 1). e HMBC
correlations from H-1ꢀꢀꢀ (ꢃ 5.31) to C-28 (ꢃ 175.2) and
was found from the combination of low concentrations of
individual agents. At 1.0 ꢇg/mL, compound 1 and acarbose
alone exhibited only 1.5% and 4.2% inhibition, respectively.
However, their combination increased the inhibition up to
32.8%. At a higher concentration of 10 ꢇg/mL, the percentage
of ꢁ-amylase inhibition of the mixture was equal to the
sum of compound 1 and acarbose, suggesting that they
produced additive inhibition. A slight increase in inhibition
was obtained from the mixture in comparison with that
achieved by the individual agents at 100 ꢇg/mL. ese find-
ings indicate that saponin 1 produced synergistic effects on
porcine pancreas ꢁ-amylase inhibition when combined with
a low concentration of acarbose.
H
C
the anomeric C-1ꢀꢀꢀ resonance shifed upfield at ꢃ 91.9
C
suggesting the galactose unit attached to C-28. e HMBC
couplings from H-1ꢀꢀꢀꢀꢀ (ꢃ 4.62) to C-2ꢀꢀꢀ (ꢃ 76.9) and from
H
H-2ꢀꢀꢀ (ꢃ 3.60) to C-1ꢀꢀꢀꢀꢀ (ꢃ 102.3) and the CCOSY crosspeak
C
betweenHH-1ꢀꢀꢀ and H-2ꢀꢀꢀ indicated that the glucose unit
attached to C-2ꢀꢀꢀ. us, compound 3 was elucidated as
3-O-{ꢀ-D-glucopyranosyl-(1→2)-[ꢀ-D-glucopyranosyl-(1→
4)]-ꢀ-D-glucuronopyranosyl} oleanolic acid 28-O-ꢀ-D-
glucopyranosyl-(1→2)-ꢀ-D-galactopyranosyl ester, named
polyscioside I.
Acarbose is a potent ꢁ-glucosidase inhibitor and has been
used in the treatment of type 2 diabetes. It can also be
used in combination with other agents, such as sulfonylurea
and metformin, in patients with diabetes. However, the
long-term use and high-dose administration of this drug
cause severe side effects, such as diarrhea, flatulence, and
abdominal or stomach pain [17–19]. Our study showed that
the combination of P. fruticosa saponin 1 with acarbose at a
low concentration produced more synergistic inhibition than
either drug alone, suggesting that they might provide a signif-
icant clinical benefit in delaying postprandial hyperglycemia
and diminishing the adverse effects of acarbose.
e inhibitory effects of the isolated compounds against
porcine pancreas ꢁ-amylase and yeast ꢁ-glucosidase were
evaluated in comparison with the antidiabetic acarbose
(Table 2). Compound 1 potently inhibited ꢁ-amylase and ꢁ-
glucosidase with IC values of 27.1 and 440.5 ꢇg/mL, respec-
50
tively. In contrast, compounds 2 and 3 did not show any effect
on both enzymes. Dou et al. reported that the glucuronic acid
unit at C-3 and glucosidic moiety at C-28 of the aglycone
were the functional groups essential for ꢁ-amylase and ꢁ-
glucosidase inhibition; the addition or replacement of glucose
by another sugar unit might thus abolish the inhibitory
activity [12]. In oral sucrose tolerance tests, several oleanolic
acid glycosides showed significant antihyperglycemic activity
in the oral sucrose tolerance in rats [13, 14]. However, the 2ꢀ-
O-ꢀ-D-glucopyranosyl unit attached to the glucuronic acid
moiety was assumed to markedly reduce the hypoglycemic
effect [15]. Consistent with this, in our study, compounds
2 and 3 possessing a 2ꢀ-O-ꢀ-D-glucopyranosyl unit were
inactive regarding ꢁ-amylase and ꢁ-glucosidase inhibition.
Previous studies showed that P. fruticosa leaf extracts strongly
inhibited rat intestinal ꢁ-glucosidase [16] and exhibited
hypoglycemic and antihyperglycemic activity in normal and
streptozotocin-diabetic albino rats [4]. Our results suggest
that the observed hypoglycemic activity might be related to
the inhibition of ꢁ-amylase and ꢁ-glucosidase enzymes.
It was of interest to establish whether compound 1
and acarbose interact synergistically or additively in the
inhibition of ꢁ-glucosidase and ꢁ-amylase. erefore, the
assay was performed in solutions containing these agents
alone or in a mixture at different concentrations. e results
showed that the combination of compound 1 and acarbose
produced a synergistic inhibitory effect on porcine pancreas
ꢁ-amylase (Figure 2) but had no effect on yeast ꢁ-glucosidase
(data not shown). Interestingly, higher synergistic inhibition
4. Conclusion
ree bisdesmosidic saponins, including two known 3-O-[ꢀ-
D-glucopyranosyl-(1→4)-ꢀ-D-glucuronopyranosyl] oleano-
lic acid 28-O-ꢀ-D-glucopyranosyl ester (1) and 3-O-{ꢀ-D-
glucopyranosyl-(1→2)-[ꢀ-D-glucopyranosyl-(1→4)]-ꢀ-D-
glucuronopyranosyl} oleanolic acid 28-O-ꢀ-D-glucopyrano-
syl ester (or polyscioside D, 2) and one new 3-O-{ꢀ-D-glu-
copyranosyl-(1→2)-[ꢀ-D-glucopyranosyl-(1→4)]-ꢀ-D-glu-
curonopyranosyl} oleanolic acid 28-O-ꢀ-D-glucopyranosyl-
(1→2)-ꢀ-D-galactopyranosyl ester (named polyscioside I,
3), were isolated from the leaves of P. fruticosa. Compound 1
showed inhibitory effects against porcine pancreas ꢁ-amylase
and yeast ꢁ-glucosidase activities and produced a synergistic
effect with acarbose in ꢁ-amylase inhibition. Further study
is needed to clarify the mechanism of enzyme inhibition as
well as the hypoglycemic properties of saponin 1.
Competing Interests
e authors declare that there are no competing interests
regarding the publication of this paper.

Journal of Chemistry
5
90
80
70
60
50
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∗
∗
40
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0
1
10
100
(ꢁg/mL)
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Compound 1
Mix
Acarbose
Figure 2: e percentage inhibition by combination of 1 and
acarbose on porcine pancreatic ꢁ-amylase. Results are expressed as
means SD; ꢈ = =. ∗ < 0.05 compared with acarbose alone.
ꢉ
Acknowledgments
is work is supported in part by the Vietnam Academy
of Science and Technology (VAST.DLT.06/14-15) and the
National Foundation for Science and Technology Develop-
ment (NAFOSTED 104.01-2011.54). e authors thank the
Institute of Chemistry (VAST) for the NMR measurements
and Dr. ML Bourguet-Kondracki (Muse´e National d’Histoire
Naturelle, Paris, France) for providing HRMS experiment.
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