Three new nortriterpene glycosides and two new triterpene glycosides from the bulbs of Scilla scilloides

Article abstract of DOI:10.1248/cpb.59.1348

Three new norlanostane-type triterpene glycosides, scillanostasides A, B, and C, and two new lanostane-type triterpene glycosides, scillanostasides D and E, were isolated from the bulbs of Scilla scilloides DRUCE (Liliaceae) along with one known norlanostane-type triterpene heptaglycoside, scillascilloside G-1. Their chemical structures were determined on the basis of spectroscopic data as well as chemical evidence.

Full text of DOI:10.1248/cpb.59.1348

1348
Regular Article
Chem. Pharm. Bull. 59(11) 1348—1354 (2011)
Vol. 59, No. 11
Three New Nortriterpene Glycosides and Two New Triterpene Glycosides
from the Bulbs of Scilla scilloides
Masateru ONO,*,a Daiki TOYOHISA,a Takahiro MORISHITA,a Hiroki HORITA,a Shin YASUDA,a
Yoichiro NISHIDA,b Toshiharu TANAKA,b Masafumi OKAWA,c Junei KINJO,c
d
Hitoshi YOSHIMITSU,d and Toshihiro NOHARA
b
a School of Agriculture, Tokai University; 5435 Minamiaso, Aso, Kumamoto 869–1404, Japan: Aso Pharmaceutical Co.,
Ltd.; 91–1 Tsukure, Kikuyo, Kumamoto 869–1101, Japan: c Faculty of Pharmaceutical Sciences, Fukuoka University;
8–19–1 Nanakuma, Jonan-ku, Fukuoka 814–0180, Japan: and d Faculty of Pharmaceutical Sciences, Sojo University;
4–22–2 Ikeda, Kumamoto 860–0082, Japan.
Received June 17, 2011; accepted August 18, 2011; published online August 23, 2011
Three new norlanostane-type triterpene glycosides, scillanostasides A, B, and C, and two new lanostane-type
triterpene glycosides, scillanostasides D and E, were isolated from the bulbs of Scilla scilloides DRUCE (Liliaceae)
along with one known norlanostane-type triterpene heptaglycoside, scillascilloside G-1. Their chemical struc-
tures were determined on the basis of spectroscopic data as well as chemical evidence.
Key words Scilla scilloides; triterpene; lanostane; glycoside; scillanostaside; Liliaceae
Scilla scilloides DRUCE is a perennial herb belonging to the the methanol (MeOH) extract of fresh bulbs of S. scilloides
Liliaceae family. The bulb of this plant has been used as a along with 13 known compounds consisting of a homostil-
foodstuff, a traditional medicine for promoting blood circula- bene, seven homoisoflavones, a xanthone, a lignan, and three
tion, an anti-inflammatory agent, and an analgesic.1) With re- norlanostane-type triterpenes. As part of an ongoing study of
gard to the chemical constituents of this bulb, the presence of this plant, we describe the isolation and structural characteri-
homoisoflavones, norlanostane-type triterpenes, and lanos- zation of three new norlanostane-type triterpene glycosides
tane-type triterpenes has been reported.2—6) In a previous and two new lanostane-type triterpene glycosides along with
paper,7) we reported the isolation and structural elucidation one known norlanostane-type triterpene glycoside from the
of a new homostilbene and two new homoisoflavones from MeOH extract.
Fig. 1. Structures of 1—8
© 2011 Pharmaceutical Society of Japan
∗ To whom correspondence should be addressed. e-mail: mono@agri.u-tokai.ac.jp

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1349
Table 1. 1H-NMR Spectral Data for Aglycone Moiety of 1 and 2 (in Pyri- Table 2. 1H-NMR Spectral Data for Sugar Moiety of 1 and 2 (in Pyridine-
dine-d5, 500 MHz)
d5, 500 MHz)
1
2
1
2
Ag-1a
1b
3.13 ddd (4.0, 4.0, 13.5)
1.83 ddd (4.0, 4.0, 13.5)
1.39 ddd (4.0, 13.5, 13.5)
2.37 m
2.09
3.62
Glc-1
4.89 d (7.5)
3.95
4.13
4.14
3.98
4.92 d (8.0)
3.96 dd (8.0, 8.5)
4.13
4.14
4.00
1.37 m
2.34 m
2.20 m
3.63
2
3
4
5
2a
2b
3
5
6a
1.87 dd (4.0, 13.0)
2.90
1.92 dd (7.0, 9.5)
2.81
6a
6b
4.50
4.20
4.57 dd (3.0, 11.0)
4.24
6b
2.86
2.77
2.50
2.29 m
2.47
1.58 m
2.67
2.06
2.72
Ara-1
5.28 d (2.0)
4.64
4.64
5.33 d (2.5)
4.69 dd (2.5, 5.5)
4.66 dd (3.0, 5.5)
4.54
4.36 dd (8.0, 11.5)
3.91 dd (3.5, 11.5)
5.20 d (8.0)
4.19
11a
11b
12a
12b
15a
15b
16a
16b
18
2
3
4
5a
3.45 d (16.0)
2.63 d (16.0)
2.67
2.08 m
2.69
4.51
4.33 dd (8.0, 11.0)
3.87
5.16 d (7.5)
4.18
4.06
4.06
5b
Glcꢂ-1
2
3
4
1.97
1.01 s
2.11
0.89 s
4.06
4.07
19
1.49 s
1.15 s
5
3.63
3.62
20
21
22
23
25
26
28
29a
29b
30
2.38 q like (6.0)
1.02 d (6.0)
5.40 d (5.5)
4.99 d (5.5)
2.52 q (7.0)
1.08 t (7.0)
1.52 s
4.43 d (11.0)
3.87 d (11.0)
1.93 s
2.41 q like (6.5)
1.11 d (6.5)
5.41 d (5.0)
5.00 d (5.0)
2.58 q (7.0)
1.08 t (7.0)
1.50 s
4.42 d (11.5)
3.83 d (11.5)
1.74 s
6a
6b
Rha-1
4.21
4.16
4.22
4.14
6.29 br s
4.81 br s
4.60 dd (3.0, 9.5)
4.25
4.84 dq (9.0, 6.5)
1.72 d (6.5)
5.02 d (7.5)
3.96
6.32 d (1.5)
4.81 dd (1.5, 3.0)
4.61 dd (3.0, 9.5)
4.27 dd (9.5, 9.5)
4.88 dq (9.5, 6.5)
1.76 d (6.5)
5.01 d (7.5)
3.98
2
3
4
5
6
Glcꢃ-1
2
COCH3
1.97 s
1.95 s
3
4
5
6a
6b
4.18
4.06
3.96
4.51
4.24 dd (9.0, 9.0)
4.07 dd (9.0, 9.0)
3.97
4.52 dd (2.0, 11.5)
4.23
d in ppm from tetramethylsilane (TMS) (coupling constants (J in Hz) are given in
parentheses).
4.23
The MeOH extract of the fresh bulbs of S. scilloides was
d in ppm from TMS (coupling constants (J in Hz) are given in parentheses).
suspended in H2O and successively extracted with ethyl ac-
etate (EtOAc) and n-butanol (BuOH). Repeated chromatog- pearance of signals due to two carbonyl groups, one acetoxy
raphy of the aqueous layer with Diaion HP20, silica gel, and group, and one oxygenated methine group and the loss of
Chromatorex octadecyl silica (ODS) column chromatogra- signals due to three methylene groups and two monosaccha-
phy as well as HPLC on ODS and silica gel led to the isola- ride units. The NMR signals were assigned in detail with the
1
tion of six compounds (1—6).
aid of H–1H correlation spectroscopy (COSY), heteronu-
Compound 6 was identified as scillascilloside G-1 on the clear multiple-quantum coherence (HMQC), heteronuclear
basis of its physical and spectral data (Fig. 1).5)
multiple-bond correlation (HMBC), and total correlation
Compound 1, tentatively named scillanostaside A, was ob- spectroscopy (TOCSY) techniques (Tables 1—4). In the
tained as an amorphous powder. In negative-ion FAB-MS, 1 HMBC spectrum of 1, the keto carbonyl carbon at d 202.0
gave an [MꢀH]ꢀ ion peak at m/z 1307 along with fragment showed long-range correlations with the methylene protons
ion peaks at m/z 1247 [1307ꢀ60 (C2H4O2, acetic acid assignable to H2-6 of aglycone moiety (Agl) at d 2.90 and
unit)]ꢀ, 1101 [1247ꢀ146 (C6H10O4, methyl pentosyl unit)]ꢀ, 2.86, with which the methine proton due to H-5 of Agl at d
1
1085 [1247ꢀ162 (C6H10O5, hexosyl unit)]ꢀ, 939 [1085ꢀ 1.87 exhibited correlations in the H–1H COSY spectrum.
146]ꢀ, and 777 [939ꢀ162]ꢀ. High-resolution (HR)-positive- Further, cross-peaks between the keto carbonyl carbon at d
ion FAB-MS showed the molecular formula of 1 to be 202.4 and the methylene protons due to H2-12 of Agl at d
C60H92O31. The 1H-NMR spectrum of 1 indicated signals due 3.45 and 2.63 were observed in the HMBC spectrum. In ad-
to four tertiary methyl groups (d 1.93, 1.52, 1.49, 1.01), two dition, the ester carbonyl carbon at d 169.8 exhibited a long-
secondary methyl groups [d 1.72 (d, Jꢁ6.5 Hz), 1.02 (d, Jꢁ range correlation with the oxygenated methine proton at d
6.0 Hz)], one primary methyl group [d 1.08 (t, Jꢁ7.0 Hz)], 5.40 assignable to H-22 of Agl. The foregoing correlations
one acetyl group (d 1.97), and five anomeric protons [d 6.29 indicated the presence of two carbonyl groups at C-7 and C-
(br s), 5.28 (d, Jꢁ2.0 Hz), 5.16 (d, Jꢁ7.5 Hz), 5.02 (d, Jꢁ 11 and an acetoxy group at C-22 of Agl. Thus, the planar
7.5 Hz), 4.89 (d, Jꢁ7.5 Hz)]. The 13C-NMR spectrum of 1 structure of 1 was a pentaglycoside of 22-acetoxy-17,23-
exhibited signals due to three keto carbonyl carbons (d epoxy-3,28-dihydroxy-27-nor-lanost-8-ene-7,11,24-trione, as
208.0, 202.4, 202.0), one ester carbonyl carbon (d 169.8), illustrated in Fig. 2. On acidic hydrolysis, 1 afforded L-rham-
two olefinic carbons (d 151.8, 151.1), and five anomeric car- nose, L-arabinose, and D-glucose, which were identified by
bons (d 106.2, 104.2, 102.5, 102.0, 101.1). The 1H- and 13C- optical rotation using chiral detection in HPLC analysis, to-
NMR spectra were similar to those of 6, apart from the ap- gether with several unidentified artificial aglycones. The cou-

1350
Vol. 59, No. 11
Table 3. 13C-NMR Spectral Data for Aglycone Moiety of 1—6 (in Pyri-
dine-d5, 125 MHz)
1
2
3
4
5
6
1
2
3
4
5
6
7
8
9
34.4
27.2
87.4
44.0
51.0
37.4
202.0
151.1
151.8
39.8
202.4
47.5
50.2
51.9
34.1
38.6
96.0
20.0
17.5
49.3
14.8
81.5
85.0
208.0
33.2
7.5
35.0
27.1
87.8
44.1
50.9
37.5
198.3
139.5
164.5
39.6
23.6
24.7
48.8
50.4
34.2
39.6
96.2
19.4
18.3
49.5
15.4
82.0
85.0
208.9
33.4
7.5
35.7
27.4
88.9
44.3
51.7
18.7
26.8
135.2
134.5
36.7
21.0
25.2
48.8
50.8
32.0
39.6
97.0
19.2
19.5
43.6
17.2
36.8
81.5
212.6
32.2
7.7
35.7
27.4
88.9
44.4
51.7
18.7
26.8
135.0
134.7
36.8
20.9
24.9
48.7
50.7
31.8
37.7
99.2
18.7
19.5
43.8
18.6
38.4
117.0
77.4
41.2
178.7
8.8
35.7
27.4
89.0
44.3
51.7
18.6
26.8
135.0
134.7
36.8
20.9
24.9
48.7
50.7
31.8
37.7
99.1
18.6
19.5
43.8
18.6
38.4
116.9
77.4
41.2
178.7
8.8
35.7
27.4
88.9
44.4
51.8
18.7
26.9
135.4
134.6
36.8
21.1
25.3
48.9
50.8
32.1
39.7
97.1
19.3
19.5
43.7
17.2
36.9
81.6
212.5
32.3
7.7
10
11
12
13
14
15
16
17
18
19
20
21
22
23
24
25
26
27
28
29
30
CO
CH3
22.9
63.1
28.6
169.8
20.7
22.5
62.9
27.5
169.9
20.8
23.0
63.1
26.3
23.1
63.1
25.9
23.0
63.1
25.9
23.1
63.1
26.4
Fig. 2. 1H–13C Long-Range Correlations Observed in the HMBC Spectra
of 1—4 (in Pyridine-d5, 500 MHz)
d in ppm from TMS.
pling constants of the signals due to the anomeric protons in
1
the H-NMR spectrum and the chemical shifts8,9) of the sig-
nals due to arabinosyl and rhamnosyl units in the 13C-NMR
spectrum suggested that all monosaccharide units were of the
pyranose form. Furthermore, the mode of glycosidic linkages
4
of glucopyranosyl units were b in C1 conformation and
those of arabinopyranosyl and rhamnopyranosyl units were a
1
in C4 conformation. The HMBC spectrum of 1 showed key
correlations between H-1 of the first glucosyl unit (Glc) and
C-3 of Agl; H-1 of the second glucosyl unit (Glcꢂ) and C-2
of the arabinosyl unit (Ara); H-1 of the rhamnosyl unit (Rha) Fig. 3. Key NOE Correlations Observed in the NOESY Spectrum of 1 (in
Pyridine-d5, 500 MHz)
and C-2 of Glcꢂ; and H-1 of the third glucosyl unit (Glcꢃ)
and C-3 of Glcꢂ (Fig. 2). In addition, the 13C-NMR spectral
data of the sugar moiety of 1 were considerably similar to L-rhamnopyranosyl-(1→2)-[O-b-D-glucopyranosyl-(1→3)]-
those of scillasaponin B (7) (Fig. 1).10) These data indicated O-b-D-glucopyranosyl-(1→2)-O-a-L-arabinopyranosyl-
that the sugar chain attached to the hydroxyl group at C-3 of (1→6)-b-D-glucopyranoside.
Agl of 1 was identical to that of 7. The stereochemistry of
Compound 2, tentatively named scillanostaside B, was ob-
Agl was defined on the basis of the nuclear Overhauser effect tained as an amorphous powder. The molecular formula of 2
spectroscopy (NOESY) and the 13C-NMR spectra of 1. In the was determined to be C60H94O30 by HR-positive-ion FAB-
NOESY spectrum, key nuclear Overhauser effect (NOE) cor- MS. The 1H-NMR spectrum of 2 was similar to that of 1, es-
relations were observed between H-3 and H3-28; H-5 and H3- pecially the signals due to the sugar moiety were almost su-
28; Hb-12 and H3-21; Ha-12 and H3-30; H3-18 and H3-19; perimposable. Furthermore, the 13C-NMR spectrum of 2 was
H3-18 and H-20; H3-19 and H2-29; H3-21 and H-22; and H3- also quite similar to that of 1, except for the loss of signal
21 and H-23 (Fig. 3). Moreover, the resonances due to C- due to one carbonyl carbon and the appearance of an addi-
20—C-26 in the 13C-NMR spectrum were superimposable on tional signal due to one methylene carbon. As in the case of
1
those of (22R,23S)-22-acetoxy-17a,23-epoxy-3b,29-dihy- 1, these H- and 13C-NMR signals were examined in detail,
droxy-27-nor-lanost-8-ene-24-one.11) The structure of 1 was and the structure of 2 was deduced to be a deoxy-derivative
thus concluded to be (22R,23S)-22-acetoxy-17a,23-epoxy- at C-11 of 1 (Fig. 2). This assumption was confirmed by the
3b,29-dihydroxy-27-nor-lanost-8-ene-7,11,24-trione 3-O-a- observation of NOE correlations similar to those of 1 in the

November 2011
1351
Table 4. 13C-NMR Spectral Data for Sugar Moiety of 1—6 (in Pyridine-
d5, 125 MHz)
gous to that of 6, showed signals due to four tertiary methyl
groups (d 1.54, 1.52, 0.94, 0.91), two secondary methyl
groups [d 1.72 (d, Jꢁ6.0 Hz), 1.04 (d, Jꢁ7.0 Hz)], one pri-
mary methyl group [d 1.07 (t, Jꢁ7.5 Hz)], and eight
anomeric protons [d 6.16 (br s), 5.26 (br s), 5.18 (d, Jꢁ
7.5 Hz), 5.16 (d, Jꢁ7.5 Hz), 5.09 (d, Jꢁ8.0 Hz), 4.96 (d,
Jꢁ7.5 Hz), 4.93 (d, Jꢁ7.5 Hz), 4.84 (d, Jꢁ7.5 Hz)] (Table 5).
The 13C-NMR spectrum of 3, which was also similar to that
of 6 with additional signals due to one pentosyl unit, con-
tained signals assignable to one keto carbonyl carbon (d
212.6), two olefinic carbons (d 135.2, 134.5), and eight
anomeric carbons (d 105.9, 105.6, 105.6, 105.2, 103.7,
102.3, 101.8, 101.1). These signals were assigned with the
help of 2D-NMR techniques as in the case of 1, and the as-
signed data of Agl were superimposable on those of 6 (Table
3). Thus, 3 was determined to be an octaglycoside of 15-de-
oxyeucosterol. On acidic hydrolysis, 3 afforded L-rhamnose,
L-arabinose, D-xylose, D-glucose, and D-galactose. Moreover,
the glycosidic linkages in glucopyranosyl, galactopyranosyl,
and xylopyranosyl units were b and those in arabinopyra-
1
2
3
4
5
6
Glc-1
106.2
75.2
78.1
72.6
75.4
68.7
101.1
77.6
71.6
66.7
62.8
102.5
76.9
88.9
69.1
77.8
61.8
102.0
72.1
72.5
74.0
69.7
18.7
104.2
74.9
78.3
71.4
78.5
62.3
106.1
75.3
78.2
72.4
75.8
68.6
101.2
77.6
71.6
66.8
62.8
102.3
76.9
89.1
69.2
77.7
61.8
102.0
72.2
72.5
74.1
69.7
18.7
104.3
74.9
78.4
71.4
78.6
62.3
105.9
75.2
78.1
72.4
75.5
68.5
101.1
77.5
71.6
66.8
62.8
102.3
76.9
88.9
69.1
77.8
61.7
101.8
72.0
72.4
73.9
69.6
18.6
103.7
73.5
88.1
69.4
77.9
62.0
105.2
71.6
83.9
69.2
76.8
61.9
105.6
75.2
77.4
71.4
76.6
70.0
105.6
74.7
78.1
70.9
66.9
106.1
75.3
78.2
72.5
75.5
68.6
101.2
77.5
71.5
66.8
62.8
102.4
76.8
89.0
69.0
77.7
61.7
101.9
72.1
72.5
74.1
69.7
18.7
103.8
73.6
88.2
69.5
77.9
62.0
105.3
71.6
84.2
69.5
76.9
62.0
106.0
75.7
78.3
71.7
78.5
62.6
106.0
75.3
78.2
72.5
75.5
68.6
101.2
77.5
71.6
66.8
62.8
102.4
77.0
88.9
69.1
77.7
61.7
101.9
72.1
72.5
74.0
69.7
18.7
103.7
73.6
88.2
69.5
77.9
62.0
105.4
71.7
84.0
69.3
76.8
62.0
105.7
75.3
77.4
71.4
76.7
70.0
105.7
74.8
78.2
71.0
67.0
106.0
75.3
78.2
72.5
75.5
68.5
101.1
77.6
71.5
66.8
62.8
102.4
76.9
89.0
69.1
77.7
61.8
101.9
72.1
72.5
74.1
69.7
18.7
103.7
73.6
88.2
69.5
77.9
62.1
105.3
71.7
84.3
69.5
76.8
62.0
106.1
75.7
78.3
71.6
78.5
62.6
2
3
4
5
6
Ara-1
2
3
4
5
Glcꢂ-1
2
3
4
5
6
Rha-1
2
3
4
5
1
nosyl and rhamnopyranosyl units were a based on the H-
and 13C-NMR spectral data. In the HMBC spectrum of 3, key
correlations were observed between H-1 of Glc and C-3 of
Agl; H-1 of Ara and C-6 of Glc; H-1 of Glcꢂ and C-2 of Ara;
H-1 of Rha and C-2 of Glcꢂ; H-1 of Glcꢃ and C-3 of Glcꢂ, H-
1 of galactosyl unit (Gal) and H-3 of Glcꢃ; H-1 of firth gluco-
syl unit (Glcꢄ) and C-3 of Gal; H-1 of xylosyl unit (Xyl) and
C-6 of Glcꢃ (Fig. 2). These data showed that for 3, one b-D-
xylopyranosyl unit may be attached to OH-6 of Glcꢄ of 6.
This was confirmed by the following evidence. Comparing
the chemical shifts of signals due to the sugar moieties be-
tween 3 and 6, glycosylation shifts12,13) were observed at C-5
and C-6 of Glcꢄ with magnitudes ꢀ1.9 and ꢅ7.4 ppm, re-
spectively, with the appearance of signals due to one terminal
xylopyranosyl unit. The resonances of other signals were al-
most identical to those of 6. In addition, the HR-positive-ion
FAB-MS of the peracetate (3a) of 3 revealed fragment ion
peaks at m/z 259.0812, 273.0966, 835.2495, and 1123.3357,
which were assigned to the fragment ions of the 2,3,4-O-tri-
acetylxylopyranosyl unit, the 2,3,4-O-triacetylrhamnopyra-
nosyl unit, the 2,3,4-O-triacetylxylopyranosyl-(1→6)-O-
(2,3,4-O-triacetyl)glucopyranosyl-(1→3)-O-(2,4,6-O-tri-
acetyl)galactopyranosyl unit, and the 2,3,4-O-triacetylxylo-
pyranosyl-(1→6)-O-(2,3,4-O-triacetyl)glucopyranosyl-
(1→3)-O-(2,4,6-O-triacetyl)galactopyranosyl-(1→3)-O-
6
Glcꢃ-1
2
3
4
5
6
Gal-1
2
3
4
5
6
Glcꢄ-1
2
3
4
5
6
Xyl-1
2
3
4
5
d in ppm from TMS.
NOESY spectrum of 2. Consequently, the structure of 2 was (2,4,6-O-triacetyl)glucopyranosyl unit, respectively, whereas
found to be (22R,23S)-22-acetoxy-17a,23-epoxy-3b,29-di- a fragment ion peak ([C14H19O9]ꢅ) generated from a 2,3,4,6-
hydroxy-27-nor-lanost-8-ene-7,24-dione 3-O-a-L-rhamnopy- tetracaetylhexosyl unit was not detected. Thus, 3 was con-
ranosyl-(1→2)-[O-b-D-glucopyranosyl-(1→3)]-O-b-D-glu- cluded to be 15-deoxyeucosterol 3-O-b-D-xylopyranosyl-
copyranosyl-(1→2)-O-a-L-arabinopyranosyl-(1→6)-b-D- (1→6)-O-b-D-glucopyranosyl-(1→3)-O-b-D-galactopyra-
glucopyranoside.
nosyl-(1→3)-O-b -D-glucopyranosyl-(1→3)-[O-a -L-
Compound 3, tentatively named scillanostaside C, was ob- rhamnopyranosyl-(1→2)]-O-b-D-glucopyranosyl-(1→2)-O-
tained as an amorphous powder. In negative-ion FAB-MS, 3 a-L-arabinopyranosyl-(1→6)-b-D-glucopyranoside.
gave an [MꢀH]ꢀ ion peak at m/z 1677 along with fragment
Compound 4, tentatively named scillanostaside D, was ob-
ion peaks at m/z 1545 [1677ꢀ132 (C5H8O4, pentosyl unit)]ꢀ, tained as an amorphous powder, and negative-ion FAB-MS
1531 [1677ꢀ146]ꢀ, 1383 [1545ꢀ162]ꢀ, 1237 [1383ꢀ showed an [MꢀH]ꢀ ion peak at m/z 1589 together with frag-
146]ꢀ, 1221 [1383ꢀ162]ꢀ, 1075 [1221ꢀ146]ꢀ, 1059 ment ion peaks at m/z 1443 [1589ꢀ146]ꢀ, 1427 [1589ꢀ
[1221ꢀ162]ꢀ, 913 [1059ꢀ146]ꢀ, 751 [913ꢀ162]ꢀ, 619 162]ꢀ, 1281 [1427ꢀ146]ꢀ, 1265 [1427ꢀ162]ꢀ, 1103
[751ꢀ132]ꢀ, and 457 [619ꢀ162]ꢀ (Fig. 4). HR-positive-ion [1265ꢀ162]ꢀ, 957 [1103ꢀ146]ꢀ, 795 [957ꢀ162]ꢀ, 663
FAB-MS showed the molecular formula of 3 to be [795ꢀ132]ꢀ, and 501 [663ꢀ162]ꢀ. The molecular formula
1
C75H122O41. The H-NMR spectrum of 3, which was analo- of 4 was analyzed as C71H114O39 using HR-positive-ion FAB-

1352
Vol. 59, No. 11
Table 5. 1H-NMR Spectral Data for Sugar Moiety of 3—6 (in Pyridine-d5, 500 MHz)
3
4
5
6
Glc-1
4.93 d (7.5)
3.96
4.18
4.19
4.94 d (8.0)
3.97
4.16
4.17
4.02
4.94 d (8.0)
3.98
4.18
4.19
3.99
4.96 d (8.0)
3.97
4.16
4.17
2
3
4
5
3.97
4.01
6a
6b
Ara-1
4.52
4.27
5.26 br s
4.66
4.53
4.26
5.28 d (2.5)
4.66
4.53
4.26
5.28 d (3.0)
4.66
4.53
4.26
5.30 br s
4.66
2
3
4.66
4.65
4.66
4.66
4
4.52
4.52
4.53
4.53
5a
5b
4.35
3.90
4.35
3.89
4.36
3.90
4.36 dd (5.0, 11.5)
3.90
Glcꢂ-1
5.16 d (7.5)
4.17
5.15 d (7.5)
4.16
5.16 d (7.5)
4.17
5.16 d (7.5)
4.18
2
3
4.04
4.05
4.03
4.06
4
4.04
4.05
4.03
4.06
5
3.59
3.57
3.58
3.59
6a
6b
4.21
4.14
4.19
4.15
4.19
4.15
4.20
4.15
Rha-1
6.16 br s
4.81 br s
4.61 dd (2.0, 9.0)
4.25
6.20 br s
4.81 br s
4.61 dd (3.0, 8.5)
4.25
4.86 dq (9.0, 6.5)
1.73 d (6.5)
4.97 d (8.0)
3.91
6.19 br s
4.81 d (2.5)
4.61 dd ((2.5, 9.0)
4.26
4.86 dq (9.0, 6.5)
1.73 d (6.5)
4.97 d (7.5)
3.91
6.21 br s
4.81 br s
4.62
4.26 dd (9.0, 9.0)
4.87 dq (9.0, 6.5)
1.74 d (6.5)
4.97 d (8.0)
3.92
2
3
4
5
4.86
6
1.72 d (6.0)
4.96 d (7.5)
3.91
4.11
3.90
Glcꢃ-1
2
3
4
5
4.11
3.91
3.91
4.11
3.90
3.90
4.11
3.90
3.91
3.91
6a
6b
Gal-1
4.41
4.09
5.09 d (8.0)
4.59 dd (8.0, 9.0)
4.23
4.42
4.08
5.12 d (7.5)
4.61 dd (7.5, 8.5)
4.21
4.40
4.09
5.08 d (7.5)
4.61 dd (7.5, 9.0)
4.24
4.43 dd (3.5, 11.5)
4.10 dd (5.0, 11.5)
5.11 d (8.0)
4.63
2
3
4.21
4
5
4.75 br d (3.0)
4.02
4.65
4.03
4.78
4.01
4.65
4.03
6a
6b
4.37
4.21
4.37
4.23
4.37
4.21
4.37 dd (3.5, 11.0)
4.23
Glcꢄ-1
5.18 d (7.5)
3.96
5.31 d (7.5)
4.00
5.21 d (7.5)
3.96
5.31 d (8.0)
4.01
2
3
4.16
4.25
4.17
4.24
4
3.99
4.19
4.01
4.18
5
3.89
3.96
3.90
3.97
6a
6b
4.76
4.10
4.50 dd (2.0, 12.0)
4.32 dd (5.5, 12.0)
4.79
4.11
4.50 dd (2.5, 11.5)
4.33 dd (5.5, 11.5)
Xyl-1
4.84 d (7.5)
3.97
4.88 d (7.5)
3.99
2
3
4.09
4.09
4
4.17
4.17
5a
5b
4.27
3.59
4.29 dd (5.0, 11.5)
3.61
d in ppm from TMS (coupling constants (J in Hz) are given in parentheses).
1
MS. The H-NMR spectrum of 4 showed signals due to four
tertiary methyl groups (d 1.54, 1.27, 0.93, 0.89), three sec-
ondary methyl groups [d 1.73 (d, Jꢁ6.5 Hz), 1.50 (d,
Jꢁ7.5 Hz), 1.09 (d, Jꢁ6.5 Hz)], and seven anomeric protons
[d 6.20 (br s), 5.31 (d, Jꢁ7.5 Hz), 5.28 (d, Jꢁ2.5 Hz), 5.15
(d, Jꢁ7.5 Hz), 5.12 (d, Jꢁ7.5 Hz), 4.97 (d, Jꢁ8.0 Hz), 4.94
(d, Jꢁ8.0 Hz)]. The 13C-NMR spectrum of 4 was analogous
to that of 6; in particular, the signals due to the sugar moiety
were almost superimposable, indicating signals due to one
carboxyl carbon (d 178.7), two olefinic carbons (d 135.0,
134.7), one acetal carbon (d 117.0), and seven anomeric car-
Fig. 4. Fragment Ions Observed in the Negative-Ion FAB-MS of 3

November 2011
1353
subjected to HPLC [Nacalai Tesque, Inc., Kyoto, Japan, Cosmosil 5SL-II,
20 mm i.d.ꢆ250 mm, CHCl3–MeOH–H2O (7 : 3 : 0.5, 6 : 4 : 1)] to give 5
(78 mg) and frs. 2.5.2.1—2.5.2.3 from fr. 2.5.2, and 3 (77 mg) from fr. 2.5.4.
HPLC (Cosmosil 5C18 AR-II, 70% MeOH) of fr. 2.5.2.2 (85 mg) afforded 4
(27 mg).
bons (d 106.1, 106.0, 105.3, 103.8, 102.4, 101.9, 101.2).
These NMR signals were assigned in detail with the aid of
2D-NMR techniques as done for 1. The assigned 13C-NMR
spectral data of Agl were quite similar to those of scilla-
saponin D (8) (Fig. 1).14) Consequently, the structure of 4
was concluded to be (23S,24S,25R)-17a,23-epoxy-24,29-di-
hydroxy-lanosta-8-en-23,26-olide 3-O-b-D-glucopyranosyl-
(1→3)-O-b-D-galactopyranosyl-(1→3)-O-b-D-glucopyra-
nosyl-(1→3)-[O-a-L-rhamnopyranosyl-(1→2)]-O-b-D-glu-
1: Amorphous powder. [a]D25 ꢀ33.2° (cꢁ4.4, pyridine). UV lmax (MeOH)
nm (log e): 264 (3.65). Positive-ion FAB-MS m/z: 1331 [MꢅNa]ꢅ. HR-posi-
tive-ion FAB-MS m/z: 1331.5508 (Calcd for C60H92O31Na: 1331.5520).
Negative-ion FAB-MS m/z: 1307 [MꢀH]ꢀ, 1247 [1307ꢀ60]ꢀ, 1161
[1307ꢀ146]ꢀ, 1146, 1101 [1247ꢀ146]ꢀ, 1085 [1247ꢀ162]ꢀ, 939
1
[1085ꢀ146]ꢀ, 777 [939ꢀ146]ꢀ, 661, 529, 367. H-NMR spectral data (in
copyranosyl-(1→2)-O-a-L-arabinopyranosyl-(1→6)-b-D- pyridine-d5, 500 MHz) d: see Tables 1 and 2. 13C-NMR spectral data: see
Tables 3 and 4.
glucopyranoside.
Compound 5, tentatively named scillanostaside E, was ob-
tained as an amorphous powder, and positive-ion FAB-MS
2: Amorphous powder. [a]D25 ꢀ33.3° (cꢁ3.0, pyridine). UV lmax (MeOH)
nm (log e): 254 (3.78). Positive-ion FAB-MS m/z: 1317 [MꢅNa]ꢅ. HR-posi-
tive-ion FAB-MS m/z: 1317.5726 (Calcd for C60H94O30Na: 1317.5725).
showed an [MꢀH]ꢀ ion peak at m/z 1721, which was 132
mass units larger than that of 4, along with fragment ion
peaks at m/z 1589 [1721ꢀ132]ꢀ, 1575 [1721ꢀ146]ꢀ, 1427
[1589ꢀ162]ꢀ, 1265 [1427ꢀ162]ꢀ, 1119 [1265ꢀ146]ꢀ,
1103 [1265ꢀ162]ꢀ, 957 [1103ꢀ146]ꢀ, 795 [957ꢀ162]ꢀ,
663 [795ꢀ132]ꢀ, and 501 [663ꢀ162]ꢀ. The molecular for-
mula of 5 was determined to be C76H122O43 by HR-positive-
Negative-ion FAB-MS m/z: 1233 [MꢀHꢀ60]ꢀ, 1087 [1233ꢀ146]ꢀ, 1071
1
[1233ꢀ162]ꢀ, 925 [1071ꢀ146]ꢀ, 763 [925ꢀ162]ꢀ, 661 [763ꢀ132]ꢀ. H-
NMR spectral data (in pyridine-d5, 500 MHz) d: see Tables 1 and 2. 13C-
NMR spectral data: see Tables 3 and 4.
3: Amorphous powder. [a]D31 ꢀ77.0° (cꢁ1.1, pyridine). Positive-ion FAB-
MS m/z: 1701 [MꢅNa]ꢅ. HR-positive-ion FAB-MS m/z: 1701.7365 (Calcd
for C75H122O41Na: 1701.7360). Negative-ion FAB-MS m/z: 1677 [MꢀH]ꢀ,
1545 [1677ꢀ132]ꢀ, 1531 [1677ꢀ146]ꢀ, 1383 [1545ꢀ162]ꢀ, 1237 [1383ꢀ
146]ꢀ, 1221 [1383ꢀ162]ꢀ, 1075 [1221ꢀ146]ꢀ, 1059 [1221ꢀ162]ꢀ, 913
[1059ꢀ146]ꢀ, 751 [913ꢀ162]ꢀ, 619 [751ꢀ132]ꢀ, 457 [619ꢀ162]ꢀ. 1H-
NMR spectral data (in pyridine-d5, 500 MHz) d: 2.58 (2H, m, H2-25 of Agl),
1.54 (3H, s, H3-28 of Agl), 1.52 (3H, s, H3-30 of Agl), 1.07 (3H, t,
Jꢁ7.5 Hz, H3-26 of Agl), 1.04 (3H, d, Jꢁ7.0 Hz, H3-21), 0.94 (3H, s, H3-19
of Agl), 0.91 (3H, s, H3-18 of Agl); sugar moiety: see Table 5. 13C-NMR
spectral data: see Tables 3 and 4.
1
ion FAB-MS. The H- and 13C-NMR spectral data of the
sugar moiety and Agl were considerably similar to those of 3
and 4, respectively. On the basis of these data, 5 was deter-
mined to be (23S,24S,25R)-17a,23-epoxy-24,29-dihydroxy-
lanosta-8-en-23,26-olide 3-O-b-D-xylopyranosyl-(1→6)-O-
b-D-glucopyranosyl-(1→3)-O-b-D-galactopyranosyl-(1→3)-
O-b-D-glucopyranosyl-(1→3)-[O-a-L-rhamnopyranosyl-
(1→2)]-O-b-D-glucopyranosyl-(1→2)-O-a-L-arabinopyra-
nosyl-(1→6)-b-D-glucopyranoside.
The study of the bulbs of S. scilloides resulted in the isola-
tion and structural elucidation of three new norlanostane-
type triterpene glycosides, named scillanostasides A, B, and
C, and two new lanostane-type triterpene glycosides, scil-
lanostasides D and E, along with one known norlanostane-
type triterpene glycoside. Among them, each scillanostasides
A and B had new aglycones; furthermore, the sugar moiety
attached to C-3 of the aglycones of scillanostasides C and E
was a new octasaccharide.
4: Amorphous powder. [a]D31 ꢀ67.3° (cꢁ1.1, pyridine). Positive-ion FAB-
MS m/z: 1613 [MꢅNa]ꢅ. HR-positive-ion FAB-MS m/z: 1613.6830 (Calcd
for C71H114O39Na: 1613.6835). Negative-ion FAB-MS m/z: 1589 [MꢀH]ꢀ,
1443 [1589ꢀ146]ꢀ, 1427 [1589ꢀ162]ꢀ, 1281 [1427ꢀ146]ꢀ, 1265 [1427ꢀ
162]ꢀ, 1119 [1265ꢀ146]ꢀ, 1103 [1265ꢀ162]ꢀ, 957 [1103ꢀ146]ꢀ, 795
[957ꢀ162]ꢀ, 663 [795ꢀ132]ꢀ, 501 [663ꢀ162]ꢀ. 1H-NMR spectral data (in
pyridine-d5, 500 MHz) d: 4.42 (1H, d, Jꢁ11.0 Hz, Ha-29 of Agl), 3.65 (1H,
d, Jꢁ11.0 Hz, Hb-29 of Agl), 3.31 (1H, dq, Jꢁ4.5, 7.5 Hz, H-25 of Agl),
2.70 (1H, d, Jꢁ14.5 Hz, Ha-22 of Agl), 2.52 (1H, dd, Jꢁ6.5, 14.5 Hz, Hb-22
of Agl), 2.20 (1H, dq-like, Jꢁ6.5, 6.5 Hz, H-20 of Agl), 1.54 (3H, s, H3-28
of Agl), 1.50 (3H, d, Jꢁ7.5 Hz, H3-27 of Agl), 1.27 (3H, s, H3-30 of Agl),
1.09 (3H, d, Jꢁ6.5 Hz, H3-21), 0.93 (3H, s, H3-19 of Agl), 0.89 (3H, s, H3-
18 of Agl); sugar moiety: see Table 5. 13C-NMR spectral data: see Tables 3
and 4.
5: Amorphous powder. [a]D31 ꢀ56.0° (cꢁ1.1, pyridine). Positive-ion FAB-
MS m/z: 1745 [MꢅNa]ꢅ. HR-positive-ion FAB-MS m/z: 1745.7228 (Calcd
for C76H122O43Na: 1745.7258). Negative-ion FAB-MS m/z: 1721 [MꢀH]ꢀ,
1589 [1721ꢀ132]ꢀ, 1575 [1721ꢀ146]ꢀ, 1427 [1589ꢀ162]ꢀ, 1265
[1427ꢀ162]ꢀ, 1119 [1265ꢀ146]ꢀ, 1103 [1265ꢀ162]ꢀ, 957 [1103ꢀ146]ꢀ,
795 [957ꢀ162]ꢀ, 663 [795ꢀ132]ꢀ, 501 [663ꢀ162]ꢀ. 1H-NMR spectral
data (in pyridine-d5, 500 MHz) d: 4.42 (1H, d, Jꢁ11.0 Hz, Ha-29 of Agl),
3.65 (1H, d, Jꢁ11.0 Hz, Hb-29 of Agl), 3.30 (1H, dq, Jꢁ4.5, 7.0 Hz, H-25
Experimental
All instruments and materials used were the same as cited in a previous
report15) unless otherwise specified.
Plant Material The bulbs of S. scilloides were cultivated in Kumamoto
prefecture, Japan, and were harvested in August 2005, and identified by one
of authors (T. Nohara). A voucher specimen has been deposited at the labo-
ratory of Natural Products Chemistry, School of Agriculture, Tokai Univer- of Agl), 2.70 (1H, d, Jꢁ14.5 Hz, Ha-22 of Agl), 2.52 (1H, dd, Jꢁ7.0,
sity.
Extraction and Isolation The crushed fresh bulbs of S. scilloides
14.5 Hz, Hb-22 of Agl), 2.20 (1H, dq-like, Jꢁ7.0, 7.0 Hz, H-20 of Agl),
1.54 (3H, s, H3-28 of Agl), 1.50 (3H, d, Jꢁ7.0 Hz, H3-27 of Agl), 1.27 (3H,
s, H3-30 of Agl), 1.09 (3H, d, Jꢁ7.0 Hz, H3-21), 0.93 (3H, s, H3-19 of Agl),
0.89 (3H, s, H3-18 of Agl); sugar moiety: see Table 5. 13C-NMR spectral
data: see Tables 1 and 2.
(18.5 kg) were extracted with MeOH at room temperature, and the solvent
was removed under reduced pressure to give a syrup (3521.7 g). The MeOH
extract was suspended in H2O and successively extracted with EtOAc and
BuOH. The aqueous layer was chromatographed over Diaion HP20 column
(H2O, MeOH, acetone) to afford MeOH eluate fraction (fr.) and acetone elu-
ate fr. The MeOH eluate fr. (76.7 g) was further subjected to Diaion HP20
column (H2O, 50% MeOH, 60% MeOH, 70% MeOH, 80% MeOH, 90%
MeOH, MeOH) to give fractions (frs.) 1—4. Fr. 2 (31.4 g) was chro-
6: Amorphous powder. [a]D16 ꢀ41.0° (cꢁ1.8, pyridine). Positive-ion FAB-
1
MS m/z: 1569 [MꢅNa]ꢅ. H-NMR spectral data (in pyridine-d5, 500 MHz)
d: 2.57 (2H, m, H2-25 of Agl), 1.55 (3H, s, H3-28 of Agl), 1.53 (3H, s, H3-
30 of Agl), 1.07 (3H, t, Jꢁ7.5 Hz, H3-26 of Agl), 1.04 (3H, d, Jꢁ6.5 Hz, H3-
21), 0.94 (3H, s, H3-19 of Agl), 0.91 (3H, s, H3-18 of Agl); sugar moiety:
matographed over silica gel [CHCl3–MeOH–H2O (8 : 2 : 0.2, 7 : 3 : 0.5, see Table 5. 13C-NMR spectral data: see Tables 3 and 4.
6 : 4 : 1, 0 : 1 : 0)] to give frs. 2.1—2.6. Fr. 2.3 (7.69 g) was subjected to Chro-
matorex ODS (60% MeOH, 70% MeOH, 80% MeOH, 90% MeOH, MeOH)
to afford frs. 2.3.1—2.3.6. HPLC (Nacalai Tesque, Inc., Kyoto, Japan, Cos-
mosil 5C18 AR-II, 20 mm i.d.ꢆ250 mm, 60% MeOH) of fr. 2.3.4 (2.63 g)
afforded 1 (42 mg), 2 (100 mg), and frs. 2.3.4.1—2.3.4.4. A part (10.0 g) of
fr. 2.4 (13.6 g) was subjected to Chromatorex ODS (70% MeOH, 80%
Acetylation of 3 Compound 3 (10 mg) in Ac2O–pyridine (1 : 1, 1 ml)
was left to stand at room temperature overnight. After removal of the reagent
under a stream of N2, the residue was partitioned between ether (1 mlꢆ3)
and H2O (1 ml). The ether layer was concentrated to afford 3a (8 mg).
3a: Amorphous powder. 1H-NMR spectral data d: 2.40 (3H, s), 2.38 (3H,
s), 2.32 (3H, s), 2.28 (3H, s), 2.22 (3H, s), 2.21 (3H, s), 2.20 (6H, s), 2.17
MeOH, 90% MeOH, MeOH) to afford 6 (8.60 g) and frs. 2.4.1—2.4.5. Fr. (3H, s), 2.13 (3H, s), 2.12 (6H, s), 2.11 (6H, s), 2.10 (6H, s), 2.08 (3H, s),
2.5 (1.68 g) was subjected to HPLC (Cosmosil 5C18 AR-II, 80% MeOH) to
2.05 (3H, s), 1.99 (3H, s), 1.98 (3H, s), 1.98 (3H, s), 1.96 (3H, s), 1.92 (3H,
give frs. 2.5.1—2.5.4. Fr. 2.5.2 (281 mg) and fr. 2.5.4 (334 mg) were each s). HR-positive-ion FAB-MS m/z: 1123.3357 (Calcd for C47H63O31:

1354
Vol. 59, No. 11
1123.3353), 835.2495 (Calcd for C35H47O23: 835.2509), 273.0966 (Calcd for
C12H17O7: 273.0974), 259.0812 (Calcd for C11H15O7: 259.0818).
(1973).
3) Kouno I., Noda N., Ida I., Sholichin M., Miyahara K., Komori T.,
Kawasaki T., Liebigs Ann. Chem., 1982, 306—314 (1982).
4) Sholichin M., Miyahara K., Kawasaki T., Heterocycles, 17, 251—257
(1982).
5) Sholichin M., Miyahara K., Kawasaki T., Chem. Pharm. Bull., 33,
1756—1759 (1985).
6) Lee S.-M., Chun H.-K., Lee C.-H., Min B.-S., Lee E.-S., Kho Y.-H.,
Chem. Pharm. Bull., 50, 1245—1249 (2002).
7) Nishida Y., Eto M., Miyashita H., Ikeda T., Yamaguchi K., Yoshimitsu
H., Nohara T., Ono M., Chem. Pharm. Bull., 56, 1022—1025 (2008).
8) Seo S., Tomita Y., Tori K., Yoshimura Y., J. Am. Chem. Soc., 100,
3331—3339 (1978).
9) Mizutani K., Hayashi A., Kasai R., Tanaka O., Yoshida N., Nakajima
T., Carbohydr. Res., 126, 177—189 (1984).
10) Mimaki Y., Ori K., Kubo S., Sashida Y., Nikaido T., Song L.-G.,
Ohmoto T., Chem. Lett., 1992, 1863—1866 (1992).
11) Amchler G., Frahm A. W., Müller-Doblies D., Müller-Doblies U., Phy-
tochemistry, 47, 429—436 (1998).
Sugar Analysis Compounds 1 (5 mg) and 3 (15 mg) were each heated
in 2 M HCl (1, 1 ml; 3, 3 ml) at a temperature of 95 °C for 1 h. The reaction
mixture was extracted with AcOEt. The aqueous layer was neutralized with
Amberlite MB-3 (Organo Co.) and then evaporated under reduced pressure
to give a monosaccharide fr. This fr. was analyzed by HPLC under the fol-
lowing conditions: column, Shodex RS-Pac DC-613, Showa Denko,
150 mmꢆ6.0 mm; solvent, CH3CN–H2O (3 : 1); flow rate, 1.0 ml/min; col-
umn temperature, 70 °C; detector, JASCO OR-2090 plus; pump, JASCO
PU-2080; and column oven, JASCO CO-2060. The retention time (tR) and
optical activity of each of the monosaccharides were detected as follows. L-
Rhamnose [tR, 3.7 min; optical activity, negative], L-arabinose [tR, 5.7 min;
optical activity, positive], and D-glucose [tR, 6.8 min; optical activity, posi-
tive] for 1; L-rhamnose [tR, 3.6 min; optical activity, negative], D-xylose [tR,
5.1 min; optical activity, positive], L-arabinose [tR, 5.7 min; optical activity,
positive], D-glucose [tR, 6.8 min; optical activity, positive], and D-galactose
[tR, 7.2 min; optical activity, positive] for 3. However, the ethyl acetate ex-
tract exhibited several spots by TLC, and the aglycones of 1 and 3 could not
be obtained.
12) Kasai R., Suzuo M., Asakawa J., Tanaka O., Tetrahedron Lett., 1977,
175—178 (1977).
Acknowledgment We express our appreciation to Mr. H. Harazono of
Fukuoka University for his measurement of the FAB-MS.
13) Tori K., Seo S., Yoshimura Y., Arita H., Tomita Y., Tetrahedron Lett.,
1977, 179—182 (1977).
14) Mimaki Y., Kubo S., Kinoshita Y., Sashida Y., Song L.-G., Nikaido T.,
Ohmoto T., Phytochemistry, 34, 791—797 (1993).
References
1) “Chuyaku Daijiten,” ed. by Koso Shin Igakuin, Shanghai Kagaku Gi- 15) Ono M., Mishima K., Yamasaki T., Masuoka C., Okawa M., Kinjo J.,
jutsu Shuppansha, 1978, p. 2270.
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