Two New Triterpene Glycosides in the Roots of Uraria crinita

Article abstract of DOI:10.1248/cpb.c18-00753

Two new triterpene glycosides, 24-deoxyoxytrogenin 3-O-α-L-rhamnopyranosyl (1→2)[β-D-glucopyra-nosyl]-β-D-galactopyranosyl (1→2)-β-D-glucuronopyranoside and sophoradiol 3-O-α-L-rhamnopyranosyl (1→2)-β-D-glucuronopyranosyl (1→2)-β-D-glucuronopyranoside with four known glycosides were isolated from a Chinese natural medicine, the roots of Uraria crinita (L.) DESV. Their structures were determined by chemical and spectral methods.

Full text of DOI:10.1248/cpb.c18-00753

Vol. 67, No. 2
Chem. Pharm. Bull. 67, 159–162 (2019)
159
Note
Two New Triterpene Glycosides in the Roots of Uraria crinita
,
a
a
a
a
a
Masafumi Okawa,* Ryo Akahoshi, Kyoko Kawasaki, Daisuke Nakano, Ryota Tsuchihashi,
a
b
Junei Kinjo, and Toshihiro Nohara
a
Faculty of Pharmaceutical Sciences, Fukuoka University; 8–19–1 Nanakuma, Jonan-ku, Fukuoka 814–0180,
b
Japan: and Faculty of Pharmaceutical Sciences, Sojo University; 4–22–1 Ikeda, Nishi-ku, Kumamoto 860–0082,
Japan.
Received September 27, 2018; accepted November 10, 2018
Two new triterpene glycosides, 24-deoxyoxytrogenin 3-O-α-L-rhamnopyranosyl (1→2)[β-D-glucopyra-
nosyl]-β-D-galactopyranosyl (1→2)-β-D-glucuronopyranoside and sophoradiol 3-O-α-L-rhamnopyranosyl
(1→2)-β-D-glucuronopyranosyl (1→2)-β-D-glucuronopyranoside with four known glycosides were isolated
from a Chinese natural medicine, the roots of Uraria crinita (L.) DESV. Their structures were determined by
chemical and spectral methods.
Key words triterpene; saponin; Uraria crinita; Leguminoseae; Fabaceae
Introduction
(HMBC)) techniques. The data of the sapogenol part were
5,8)
Uraria crinita (L.) DESV. is distributed in southern Japan, identical to those of 24-deoxyoxytrogenin
except for C-2
9,10)
Taiwan, South China, and India. Its aerial parts and roots have and -3 signals due to glycosylation.
Moreover, the H-1 (d,
been used in traditional Chinese medicine for the treatment of J=7.9Hz at δ 4.98) of glucuronic acid correlated with the
kidney disease as well as its anti-inflammatory and antitoxic C-3 (δ 90.8) of that in the HMBC (Fig. 1). Thus, 1 was char-
effects. Several flavone C-glycosides were previously isolated acterized as 24-deoxyoxytrogenin 3-O-α-L-rhamnopyranosyl
1)
from the aerial parts of this species, while some triterpenes, (1→2)[β-D-glucopyranosyl]-β-D-galactopyranosyl (1→2)-β-D-
2)
isoflavonoids, and flavonolignans were found in its roots. In glucuronopyranoside.
3,4)
our search for bioactive substances in leguminous plants,
Compound 2 was obtained as an amorphous powder show-
we have studied the constituents of this plant. This paper deals ing [α]D −44.8° [pyridine–H O=(1:1)]. The negative FAB-
2
−
with the isolation and structural elucidation of triterpene sa- MS exhibited a peak at m/z 939 due to [M−H] and fragment
ponins from its roots. Specifically, the methanol extract of the peaks at m/z 793 [M−H−Rha] and 599 [M−H−Rha−GlcA].
roots of U. crinita was separated by normal and reverse-phase The exact measurement under HR conditions showed that the
−
column chromatography to give two new triterpene glycosides composition is C H O at m/z 939.4976 [M−H] in the HR/
4
8
75 18
(
1 and 2) along with four known ones (3–6) (Chart 1). Com- negative FAB-MS. The monosaccharide mixture obtained by
pounds 3–6 were identified as abrisaponin So , abrisaponin F, acid hydrolysis of 2 revealed the presence of D-glucuronic
1
7
)
1
phaseoside IV, and kaikasaponin III, respectively, according to acid and L-rhamnose. The H-NMR spectrum showed eight
the reported saponin data.
Compound 1 was obtained as a brown amorphous powder
5,6)
tertiary methyl signals between δ 0.84 and 1.42. Moreover the
H-signals displayed the presence of three anomeric protons at
1
showing [α] −128.4° [c=0.5, pyridine–H O=(1:1)]. The ne- δ 5.07 (1H, d, J=7.6Hz), δ 5.82 (1H, d, J=7.3Hz), and δ 6.31
D
2
−
13
gative FAB-MS exhibited a peak at m/z 1117 due to [M−H] . (1H, d, J=1.5Hz). The C-NMR spectrum of 2 exhibited 48
The exact measurement under high-resolution (HR) conditions signals consisting of a triterpene moiety and three sugar moi-
showed that the composition is C H O at m/z 1117.5413 eties. The data of the sapogenol part, including two oxygen-
5
4
85 24
−
[
M−H] in the HR/negative FAB-MS. The monosaccharide bearing carbons (δ 90.7 and 75.9), were superimposable on
6)
mixture obtained by acid hydrolysis of 1 revealed the presence those of abrisaponin So1. Moreover, the signals due to sugar
11)
of D-glucuronic acid, D-galactose, D-glucose, and L-rhamnose signals at C-3 were identical to those of yunganoside N2.
by the following analytical method of Tanaka et al. The
H-NMR spectrum showed seven tertiary methyl signals be- the H-1 (d, J=7.6Hz, δ 5.07) of glucuronic acid and the C-3
tween δ 0.88 and 1.76. Moreover, the H-signals displayed the (δ 90.7) of the aglycone, between C-2 (δ 79.3) of glucuronic
7
)
In the HMBC, the correlations were observed between
1
1
presence of four anomeric protons at δ 4.98 (1H, d, J=7.9Hz, acid and H-1 (d, J=7.3Hz, δ 5.82) of the inner glucuronic
glcA H-1), 5.00 (1H, d, J=7.6Hz, glc H-1), 5.59 (1H, d, acid, and between C-2 (δ 78.6) of the inner glucuronic acid
J=7.9Hz, gal H-1) and 6.17 (1H, d, J=1.5Hz, rha H-1). The and H-1 (d, J=1.5Hz, δ 6.31) of the terminal rhamnose
13
C-NMR spectrum of 1 exhibited 54 signals indicating a (Fig. 2). Thus, 2 was characterized as sophoradiol 3-O-α-L-
triterpene moiety, including two oxygen-bearing carbons (δ rhamnopyranosyl (1→2)-β-D-glucuronopyranosyl (1→2)-β-D-
5.7 and δ 90.8) and a carboxylic carbon (δ 182.0). The carbon glucuronopyranoside.
signals due to the sugar part were in agreement with those of These saponins were characteristic glycosides obtained
arbisaponin F. The structure of the sapogenol part was veri- from Leguminoseae plants.
7
6
)
1
1
fied by various two-dimensional (2D) NMR ( H– H correla-
tion spectroscopy (COSY), heteronuclear multiple quantum Experimental
coherence (HMQC), heteronuclear multiple bond connectivity
General Procedures Optical rotations were determined
*
To whom correspondence should be addressed. e-mail: mokawa@fukuoka-u.ac.jp
©
2019 The Pharmaceutical Society of Japan

1
60
Chem. Pharm. Bull.
Vol. 67, No. 2 (2019)
Chart 1
Fig. 1. NOE and Key HMBC Correlations of Compound 1
on a JASCO DIP-360 polarimeter (l=0.5). FAB-MS data were
Fig. 2. Key HMBC Correlations of Compound 2
obtained in a glycerol matrix in positive ion mode using a
1
JEOL JMS-HX110 mass spectrometer. H-NMR (500MHz)
13
and C-NMR (125MHz) spectra were obtained with a JEOL saponin fraction (2.4g) by eluting with 80% MeOH. This
JNM-A500 spectrometer. Column chromatography was car- saponin fraction was then subjected to MCI gel CHP 20P col-
ried out with silica gel 60 (230–400 mesh and ASTM; Kanto umn chromatography using 60% MeOH→MeOH to give five
Kagaku, Japan), Diaion HP-20 (Mitsubishi Kasei, Japan), fractions. Fraction 1 (129mg) was further chromatographed
Sephadex LH-20 (25–100nm; Pharmacia Fine Chemicals, on octadecyl silica (ODS) (70% MeOH +0.2% AcOH) and
Japan), MCI gel CHP 20P (75–150nm; Mitsubishi Kasei), and silica gel (BuOH–AcOH–water=4:1:2) to provide compound
COSMOSIL (75C -OPN and 150C -OPN; Nacalai Tesque 1 (11.9mg). Fraction 2 (139mg) was also chromatographed
18
18
Inc., Japan). TLC was conducted on a precoated silica-gel on ODS (70% MeOH +0.2% AcOH) and silica gel (CHCl3–
0F254 plate (0.2mm; Merck, Germany), and detection was MeOH–water=7:3:0.5+0.2% AcOH) to afford compound 2
achieved by spraying it with 10% H SO followed by heating. (16.2mg). Fractions 3 (313mg) and 4 (320mg) were chromato-
6
2
4,
Plant Material Uraria crinita (L.) DESV. (Leguminosae) graphed on ODS (30% CH CN) and silica gel (CHCl –MeOH–
3
3
was cultured at the Botanical Garden of Fukuoka University water=7:3:0.5+0.2% AcOH/BuOH–AcOH–water=4:1:5)
in 2013, and its roots were collected. These seeds were pur- to provide compounds 3 (176.3mg), 4 (18.7mg), 5 (7.4mg),
chased from Yoshioka Seed Bank (Nagano, Japan).
Extraction and Isolation The fresh roots (485g) of Ura-
and 6 (17.7mg).
Compound
2
0
1
An amorphous powder. [α]D −128.4°
ria crinita were extracted with MeOH. This MeOH extract [c=0.5, pyridine–H O: (1:1)]. HR FAB-MS m/z 1117.5413
2
(
15.2g) was subjected to Diaion HP-20 to remove the sugar (Calcd. for C H O : 1117.5431). Negative FAB-MS m/z 1117
54 85 24
−
part, eluting first with water and then with MeOH. The MeOH [M-H] .
portion (5.5g) was applied to Sephadex LH-20 to collect a
1
H-NMR (pyridine-d ) δ: Table 1.
5

Vol. 67, No. 2 (2019)
Chem. Pharm. Bull.
161
1
3
20
C-NMR: Tables 2, 3.
Compound 2 An amorphous powder. [α]D −44.8°[c=0.5,
Identification of Sugars for 1 A solution of 1 (5.0mg) in pyridine–H O (1:1)]. HR FAB-MS m/z 939.497 (Calcd for
.5M HCl (1mL) was heated at 95°C in screw-capped vials C H O : 939.4953). Neg. FAB-MS m/z 939 [M−H] , 793
for two hours. The mixture was evaporated to dryness in [M−H−Rha] , 599 [M−H−Rha−GlcA] .
vacuo, dissolved in 100µL of L-cysteine solution (10mg/mL
pyridine), and reacted at 60°C for one hour. A solution (1µL)
of o-toryl isothiocyanate was added to the mixture, followed
2
−
0
4
8
75 18
−
−
1
H-NMR (pyridine-d ) δ: Table 1.
5
13
C-NMR: Tables 2, 3.
Identification of Sugars for 2 In the same manner as de-
by heating at 60°C for one hour. The final mixture was direct- scribed above for 1, the t values of the peaks of D-glucuronic
R
ly analyzed by HPLC [COSMOSIL 5C18 AR II (250×4.6mm acid (18.958min) and L-rhamnose (31.294min) were confirmed
i.d.; Nacalai Tesque Inc.); mobile phase CH CN in 50mM in those of comprised sugar of 2 at 18.659 (D-glucuronic acid)
3
H PO (4–30% linear gradient at 0–39min and 30%–75% lin- min and 31.744 (L-rhamnose) min.
3
4
ear gradient at 39–54min); flow rate, 0.8mL/min; and column
temperature, 35°C ; detection, 250nm]. The tR values of the der. [α]D −2.7° [c=0.5, pyridine–H O (1:1)]. Neg. FAB-MS
peaks of D-galactose (16.008min), L-galactose (16.561min), D- m/z 1087 [M–H] . H-NMR (pyridine-d ) δ: 0.87, 0.99, 1.00,
Compound 3 (Abrisaponin So ) A white amorphous pow-
1
2
0
2
−
1
5
glucose (18.564min), L-glucose (16.960min), D-glucuronic acid 1.22, 1.24, 1.27, 1.28, 1.40 (each 3H, s, tert-Me×8), 1.78 (3H,
(18.958min), and L-rhamnose (31.294) were confirmed in those d, J=6.1Hz, rha H-6), 5.01 (1H, d, J=7.6Hz, glcA H-1), 5.02
of comprised sugars of 1 at 16.215 (D-galactose), 18.602 (D- (1H, d, J=7.3Hz, glc H-1), 5.32 (1H, brs, H-12), 5.64 (1H, d,
glucose), 19.353 (D-glucuronic acid), and 31.596 (L-rhamnose) J=7.6Hz, gal H-1), 6.21 (1H, s, rha H-1).
13
minutes.
C-NMR: Tables 2, 3.
1
Table 1. H-NMR Data for Compounds 1 and 2
1
2
1
2
1
2
α1.04 (o)
α0.88 (t-like, 13.7)
β1.44 (o)
Glc A
Glc A’
Gal
1
2
3
4
5
6
1
2
3
4
5
6
1
2
3
4
5
6
5.00 (d, 7.6)
4.52 (t, 9.2)
4.55 (o)
5.07 (d, 7.6)
4.50 (o)
β1.46 (o)
α1.94 (o)
α1.86 (o)
4.35 (o)
β2.22 (br d 9.8)
3.32 (dd, 11.6, 4.6)
β2.17 (o)
4.40 (o)
4.43 (t, 9.7)
4.63 (o)
3
4
5
6
3.34 (dd, 11.6, 4.3)
4.59 (d, 9.2)
0.97 (br d, 11.3)
α1.60 (o)
0.8.1 (br d, 11.3)
α1.33 (o)
5.82 (d, 7.3)
4.34 (o)
β1.80 (o)
β1.54 (o)
4.27 (t, 8.9)
4.50 (o)
7
α1.34 (br d, 11.3)
β1.54 (br d, 11.3)
α1.34 (o)
β1.54 (o)
4.63 (o)
8
9
0
1
2
3
4
5
1.61 (o)
1.61 (o)
5.59 (d, 7.9)
4.57 (o)
1
1
1
1
1
1
1.85 (o)
1.83 (o)
4.08 (dd, 8.5, 5.2)
4.71 (o)
5.37 (br s)
5.31 (brs)
3.94 (t, 7.9)
4.18 (o)
α1.02 (o)
β1.80 (o)
α1.22 (o)
β1.36 (o)
α1.03 (dd, 12.5, 5.8)
β1.85 (o)
4.26 (o)
Glc
1
2
3
4
5
6
4.98 (d, 7.6)
3.93 (o)
1
6
α1.43 (o)
β1.87 (o)
3.89 (m)
1
1
1
7
8
9
4.03 (t, 9.5)
4.19 (o)
2.46 (br d, 13.3)
α1.62 (o)
2.35 (br d, 13.1)
α1.18 (br d, 12.2)
β1.88 (o)
4.30 (t, 9.5)
4.42 (o)
β2.73 (t, 13.3)
2
2
0
1
Rha
1
2
3
4
5
6
6.16 (d, 1.5)
4.82 (dd, 3.4, 1.5)
4.71(o)
6.31 (br s)
α2.53 (br d, 13.3)
β2.11 (dd, 13.4, 10.4)
3.99 (o)
α1.77 (t-like, 7.9)
β1.63 (o)
3.3.75 (dd, 7.2, 3.6)
1.42 (s)
4.73 (br s)
4.70 (dd, 9.4, 3.2)
4.36 (o)
2
2
2
2
2
2
2
2
3
2
3
4
5
6
7
8
9
0
4.28 (td, 9.5, 4.6)
5.00 (o)
1.37 (s)
5.00 (dd, 9.4, 6.4)
1.81 (m)
1.20 (s)
1.21 (s)
1.75 (d, 6.4)
0.88 (s)
0.84 (s)
0.99 (s)
0.98 (s)
1.26 (s)
1.26 (s)
1.25 (s)
1.21 (s)
0.98 (s)
1.76 (s)
1.24(s)
Chemical sifts (δ) are shown in ppm, (o) means overlapping signal.

1
62
Chem. Pharm. Bull.
Vol. 67, No. 2 (2019)
C-NMR Data for Compounds 1–6 (Sugar Moieties)
13
13
Table 2.
C-NMR Data for Compounds 1–6 (Aglycone Moieties)
Table 3.
1
2
3
4
5
6
1
2
3
4
5
6
C-1
2
39.3
26.6
90.8
40.1
56.3
18.9
33.6
40.3
48.2
37.2
24.2
39.2
26.8
90.7
40.0
56.3
18.8
33.6
40.3
48.3
37.2
24.1
122.9
145.1
42.8
26.7
29.2
38.3
45.8
47.1
31.1
42.3
75.9
28.7
17.2
16.0
17.5
25.9
21.2
33.2
29.0
39.1
26.6
90.6
40.0
56.1
18.7
33.4
40.2
48.1
37.0
24.0
122.8
145.0
42.6
26.7
29.0
38.2
45.7
46.9
31.0
42.3
75.8
28.6
17.0
15.9
17.4
25.8
21.2
33.3
28.9
39.2
27.7
90.8
40.1
56.3
18.8
33.3
40.3
48.3
37.2
24.1
124.3
142.2
42.4
27.7
27.7
48.3
48.2
47.0
34.5
51.3
217.4
28.7
17.1
15.9
17.3
25.8
21.3
32.2
25.7
39.2
26.7
90.6
40.0
56.2
18.7
33.2
40.2
48.3
37.1
24.1
124.3
142.1
42.7
26.6
27.7
48.2
48.1
46.9
34.4
51.2
217.1
28.6
17.0
15.9
17.2
25.8
21.2
32.1
25.6
39.1
26.5
90.6
39.9
56.1
18.6
33.4
40.1
48.1
37.0
24.0
122.7
144.9
42.6
26.5
29.0
38.1
45.6
46.9
31.0
42.3
75.7
28.8
16.9
15.8
17.3
25.8
21.2
33.2
28.5
Glc A C-1
105.6
78.7
78.7
73.8
77.2
173.7
105.4
79.3
77.4
73.6
77.4
173.1
102.8
78.6
79.0
73.6
77.2
172.8
105.7
78.6
76.3
73.7
77.3
173.3
105.7
78.6
76.3
73.4
78.4
172.4
105.6
79.1
76.7
73.6
77.2
173.4
105.2
79.0
76.6
73.9
76.6
—
2
3
3
4
4
5
5
6
6
7
Glc A′ C-1
8
2
3
4
5
6
9
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
123.7
144.5
42.8
26.8
29.2
38.2
45.0
41.7
42.9
37.7
75.7
28.7
17.1
16.0
17.5
25.8
21.1
182.2
25.2
Gal
Glc
Rha
C-1
2
102.6
76.4
85.4
70.0
75.7
62.5
105.7
75.7
78.4
71.7
78.3
62.6
102.2
72.6
72.4
74.5
69.5
19.1
102.6
76.3
85.2
70.0
75.8
62.4
105.7
75.8
78.3
71.6
78.3
62.5
102.2
72.6
72.4
74.4
69.4
19.1
102.2
77.0
85.4
69.5
75.6
62.4
105.8
75.0
78.3
71.6
78.3
62.6
102.6
72.4
72.6
74.4
69.9
19.1
102.2
78.7
75.8
70.7
76.2
62.4
101.9
78.8
76.5
70.7
76.1
62.4
3
4
5
6
C-1
2
3
4
5
6
C-1
2
102.3
72.4
72.6
74.4
69.7
19.1
102.9
72.5
72.6
74.4
69.6
19.0
102.7
72.4
72.4
74.3
69.4
18.9
3
4
5
6
Compound 4 (Abrisaponin F) A white amorphous pow- References
2
0
der. [α] −32.1° (c=0.5, pyridine). Neg. FAB-MS m/z 1085
1) Morita N., Arisawa M., Nagase M., Hsu H.-Y., Chen Y.-P., Yakugaku
Zasshi, 97, 701–703 (1977).
D
−
1
[
M−H] . H-NMR (pyridine-d ) δ: 0.88 (6H, s, tert.-Me×2),
5
2)
Wang Y.-Y., Zhang X.-Q., Gong L.-M., Ruan H.-L., Pi H.-F., Zhang
0
(
5
.93, 1.00, 1.17, 1.18, 1.31, 1.40 (each 3H, s, tert.-Me×6), 1.76
3H, d, J=6.1Hz, rha H-6), 4.99 (1H, d, J=7.6Hz, glcA H-1),
.02 (1H, d, J=7.3Hz, glc H-1), 5.45 (1H, brs, H-12), 5.61
1H, d, J=7.6Hz, gal H-1), 6.18 (1H, d, J=1.5Hz, rha H-1).
Y.-H., Zhongguo Yaoxue Zhazhi, 44, 1217–1220 (2009).
Kinjo J., Nohara T., “Towards Natural Medicine Research in the 21
st
3)
Century”, ed. by Ageta H., Aimi N., Ebizuka Y., Fujita T., Honda
G., Elsevier, Tokyo, 1998, pp. 237–249.
(
13
C-NMR: Tables 2, 3.
Compound 5 (Phaseoside IV) A white amorphous pow-
4)
Kinjo J., Nohara T., “Studies in Natural Products Chemistry”, ed.
by Atta-ur-Rahman, Vol. 25, Elsevier Science B. V., 2001, pp.
89–124.
2
0
der. [α] −28.6° (c=0.5, pyridine) Neg. FAB-MS m/z 923
D
−
1
[
M–H] . H-NMR (pyridine-d ) δ: 0.86, 0.86, 0.92, 0.96,
5) Miyao H., Sakai Y., Takeshita T., Kinjo J., Nohara T., Chem.
Pharm. Bull., 44, 1222–1227 (1996).
5
1
.17, 1.18, 1.28, 1.42 (each 3H, s, tert.-Me×8), 1.76 (3H, d,
J=6.1Hz, rha H-6), 5.06 (1H, d, J=7.0Hz, glcA H-1), 5.70
1H, d, J=7.3Hz, gal H-1), 6.29 (1H, s, rha H-1).
C-NMR: Tables 2, 3.
Compound 6 (Kaikasaponin III) A white amorphous
powder. [α] −14.9° (c=0.5, pyridine). Neg. FAB-MS m/z
25 [M–H] . H-NMR (pyridine-d ) δ: 0.87, 1.00, 1.00, 1.20,
.23, 1.28, 1.28, 1.37 (each 3H, s, tert.-Me×8), 1.74 (3H, d,
J=5.5Hz, rha H-6), 5.57 (1H, d, J=5.7Hz, gal H-1), 6.20
1H, s, rha H-1).
6
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(
13
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75–178 (1977).
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2
0
D
2
−
1
9
5
9)
1
1
1
(
Lett., 18, 179–182 (1977).
11) Ji S., Wang Q., Qiao X., Guo H.-C., Yang Y.-F., Bo T., Xiang C.,
Guo D.-A., Ye M., J. Pharm. Biomed. Anal., 90, 15–26 (2014).
13
C-NMR: Tables 2, 3.
Conflict of Interest The authors declare no conflict of
interest.