Acylated glycosidic acid methyl esters generated from the convolvulin fraction of Rhizoma Jalapae Braziliensis by treatment with indium(III) chloride in methanol

Article abstract of DOI:10.1248/cpb.c16-00673

Four hexaglycosides of methyl 3S,12S-dihydroxyhexadecanoate (1-4) were provided after treatment of the crude convolvulin fraction from Rhizoma Jalapae Braziliensis (the root of Ipomoea operculata (GOMES) MART., Convolvulaceae) with indium(III) chloride in

Full text of DOI:10.1248/cpb.c16-00673

Vol. 65, No. 1
Chem. Pharm. Bull. 65, 107–111 (2017)
107
Note
Acylated Glycosidic Acid Methyl Esters Generated from the Convolvulin
Fraction of Rhizoma Jalapae Braziliensis by Treatment with Indium(III)
Chloride in Methanol
,a
Masateru Ono,* Satoko Oda,a Shin Yasuda,a Tomoko Mineno,b Masafumi Okawa,c Junei Kinjo,c
Hiroyuki Miyashita,d Hitoshi Yoshimitsu,d Toshihiro Nohara,d and Kazumoto Miyaharae
b
a School of Agriculture, Tokai University; 9–1–1 Toroku, Higashi-ku, Kumamoto 862–8652, Japan: Faculty of
c
Pharmacy, Takasaki University of Health and Welfare; 60 Nakaorui, Takasaki, Gunma 370–0033, Japan: Faculty
d
of Pharmaceutical Sciences, Fukuoka University; 8–19–1 Nanakuma, Jonan-ku, Fukuoka 814–0180, Japan: Faculty
e
of Pharmaceutical Sciences, Sojo University; 4–22–2 Ikeda, Nishi-ku, Kumamoto 860–0082, Japan: and Faculty of
Pharmaceutical Sciences, Setsunan University; 45–1 Nagaotoge-cho, Hirakata, Osaka 573–0101, Japan.
Received August 24, 2016; accepted October 21, 2016
Four hexaglycosides of methyl 3S,12S-dihydroxyhexadecanoate (1–4) were provided after treatment of the
crude convolvulin fraction from Rhizoma Jalapae Braziliensis (the root of Ipomoea operculata (GOMES) MART.,
Convolvulaceae) with indium(III) chloride in methanol. The structures of 1–4 were elucidated on the basis of
spectroscopic and chemical methods. Their sugar moieties were partially acylated with organic acids includ-
ing (3S,9R)-3,6:6,9-diepoxydecanoic (exogonic) acid, (E)-2-methylbut-2-enoic (tiglic) acid, and isovaleric acid.
Key words resin glycoside; convolvulin; Ipomoea operculata; Rhizoma Jalapae Braziliensis; acylated glyco-
sidic acid methyl ester; indium(III) chloride
Rhizoma Jalapae Braziliensis, the root of Ipomoea opercu- indium(III) chloride in MeOH, as in the cases of the crude
lata (GOMES) MART. (Convolvulaceae), is used as a crude laxa- convolvulin fractions from Pharbitis Semen11) and seeds of
tive drug. Its active ingredients are known to be ether-soluble Quamolit pennata.12) The treated fraction was successively
resin glycoside called jalapin and ether-insoluble resin glyco- separated over Sephadex LH-20 and silica gel column chroma-
side called convolvulin.1,2) In preceding papers, we reported tography and HPLC on octadecyl silica (ODS), affording four
seven glycosidic acids, operculinic acids A–G, and two or- compounds, referred to as IOM-1–IOM-4.
ganic acids, n-decanoic acid and n-dodecanoic acid, as the al-
IOM-1 (1) was obtained as an amorphous powder and its
kaline hydrolysis products of the crude jalapin fraction of the molecular formula was determined as C73H122O37 by high-res-
crude drug, and isolation and structural elucidation of 18 new olution (HR)-positive-ion FAB-MS at m/z 1613.7570. Alkaline
genuine jalapins, operculins I–XVIII, from the fraction.3–8) We hydrolysis of 1 afforded an organic acid fraction and a glyco-
also reported a glycosidic acid, operculinic acid H, along with sidic acid. The organic acid fraction was subjected to GC to
three organic acids such as isovaleric acid, (E)-2-methylbut- reveal the presence of isovaleric acid, tiglic acid, and exogonic
2-enoic (tiglic) acid, and (3S,9R)-3,6:6,9-diepoxydecanoic acid. The glycosidic acid was identified as operculinic acid H
1
1
(exogonic) acid, which exists as an equilibrium mixture (ca. (5) based on its H-NMR spectral data.9) The H-NMR spec-
1:1) at the C-6 spiro center, formed by the alkaline hydrolysis trum of 1 showed signals due to one each of isovaleroyl residue
of the crude convolvulin fraction.9) As part of our studies on (Iva), tigloyl residue (Tig), exogonoyl residue (Exg), and methyl
the resin glycosides from this crude drug, herein we report ester (5a)9) of 5. Moreover, the spectrum showed that 1 was a
the isolation and structural elucidation of four acylated glyco- mixture of epimers that differ in the spirocyclic carbon configu-
sidic acid methyl esters. We could afford these methyl esters ration only at C-6 of Exg in a ratio of approximately 1:1 (Table
by effectively treating the crude convolvulin fraction with 1). We thus envisioned the isolation of these epimers. The
indium(III) chloride in methanol (MeOH), a catalyst reagent HPLC on naphthylethyl group bonded silica (π-nap) of 1 yielded
1
for the mild methyl esterification of carboxylic acids, as previ- two compounds (1a, b). However, the H-NMR spectra of 1a
ously reported.10)
and b were quite similar to that of 1, and further each HPLC
analysis using the π-nap column of 1a and b afforded two peaks
in which the Rt values matched with those of 1a and b, respec-
Results and Discussion
Powdered roots of I. operculata, which was purchased from tively. Because the data show that the interconversion of the
Paul Müggenburg Gmbh & Co., a botanical raw materials epimers of Exg is facile, the structure analyses were carried out
company in Hannover, Germany, were extracted with MeOH. using a mixture of the epimers. The facile epimerization at C-6
This extract was suspended in H2O, and then successively of exogonic acid was also reported by Lawson et al.13)
1
extracted with ether and n-butanol (BuOH). The n-BuOH-
Assignments of the H- and 13C-NMR signals of 1 were
soluble fraction was subjected to MCI gel CHP20 column conducted with the aid of the 1H–1H correlation spectros-
chromatography to yield a crude convolvulin fraction.3) De- copy (COSY), heteronuclear multiple-quantum coherence
spite numerous trials, our attempt for the isolation of pure (HMQC), heteronuclear multiple-bond correlation (HMBC),
resin glycosides from the crude convolvulin fraction has never and 1H–1H total correlation spectroscopy (TOCSY) spectra
1
become successful. This fraction was therefore treated with (Tables 1, 2). Comparing the chemical shifts of the H-NMR
*To whom correspondence should be addressed. e-mail: mono@agri.u-tokai.ac.jp
© 2017 The Pharmaceutical Society of Japan

108
Chem. Pharm. Bull.
Vol. 65, No. 1 (2017)
Table 1. 1H-NMR Spectral Data for 1 and 2 (in Pyridine-d5, 500MHz)
signals due to the presence of sugar moieties between 1 and
5a, significant downfield shifts owing to acylation were ob-
served for the signals because of H-2 of the third glucosyl
unit (Glc″), H-2 of the second rhamnosyl unit (Rha′), and
H-4 of Rha′ of 1. From these data, the locations of ester link-
ages were concluded to be at OH-2 of Glc″, OH-2 of Rha′,
and OH-4 of Rha′. The sites of each ester linkage of Iva,
Tig, and Exg were corroborated from the HMBC spectrum
of 1, that is, key cross-peaks between H-2 of Glc″ and C-1
of Iva, H-2 of Rha′ and C-1 of Exg, H-4 of Rha′ and C-1
of Tig, and methoxy protons and C-1 of the aglycone moi-
ety (Agl) were observed (Fig. 1). Therefore, Iva, Tig, and
Exg were located at OH-2 of Glc′, OH-2 of Rha′, and OH-4
of Rha′, respectively. Consequently, the structure of 1 was
defined as methyl 3S,12S-dihydroxyhexadecanoate 12-O-β-
Position
1
2
Glc-1
4.83 d (7.5)
4.30a)
4.90 d (7.5)
4.28a)
2
3
4.43a)
3.86a)
3.96a)
4.50a)
4
3.87 dd (9.0, 9.0)
4.00a)
5
6
4.63 brd (11.0)
4.05a)
4.52a)
4.02a)
6
Glc′-1
5.75 d (8.0)
4.10a)
5.71 d (7.5)
4.16a)
2
3
3.95a)
3.99 dd (8.0, 9.0)
3.87 dd (9.0, 9.0)
3.71 m
4
3.81 dd (9.0, 9.0)
3.70 m
5
6
4.33 dd (2.0, 11.0)
4.12a)
4.32 dd (2.0, 12.0)
4.15a)
6
D-glucopyranosyl-(1→3)-O-(2-O-(3S,9R)-exogonoyl,
4-O-
Glc″-1
5.11 d (7.5)
5.15 d (8.0)
tigloyl)-α-L-rhamnopyranosyl-(1→2)-[O-(2-O-isovaleroyl)-β-D-
glucopyranosyl-(1→3)]-O-β-D-glucopyranosyl-(1→2)-[O-α-L-
rhamnopyranosyl-(1→6)]-O-β-D-glucopyranoside, as shown in
Fig. 2.
2
5.52 dd (7.5, 9.5), 5.51 dd (7.5, 9.5) 5.54 dd (8.0, 9.5), 5.54 dd (8.0, 9.5)
3
4.41a)
4.08a)
4.08a)
4.53a)
4.25a)
4.44a)
4.12a)
4.14a)
4.52a)
4.27a)
4
5
6
IOM-2 (2) was obtained as an amorphous powder and af-
forded isovaleric acid, exogonic acid, and 5 upon alkaline
hydrolysis. Its positive-ion FAB-MS exhibited an [M+Na]+
ion peak at m/z 1531, 82 mass units (tigloyl residue) less than
the corresponding peak of 1. The molecular formula of 2
was established as C68H116O36 by HR-positive-ion FAB-MS.
6
Glc‴-1
5.22 d (7.5)
3.88a)
4.03a)
3.90a)
4.12a)
4.51a)
4.13a)
5.49 d (8.0)
4.06a)
4.16a)
4.04a)
4.02a)
4.48a)
4.19a)
2
3
4
5
1
The H- and 13C-NMR spectra of 2 were similar to those of
6
6
1 except for the loss of signals for Tig in 1 (Tables 1, 2). The
Rha-1
5.44 s
5.41 s
4.52a)
1
chemical shifts of the H-NMR signals owing to the presence
2
4.55 d (3.5)
4.48 dd (3.5, 9.0)
4.25 dd (9.0, 9.0)
4.31a)
of sugar moieties in 2 and 1 were compared. The signal due
to H-4 of Rha′ of 2 showed remarkable upfield shift, whereas
the signals of H-2 of Glc″ and H-2 of Rha′ appeared at posi-
tions similar to those of 1. The above data suggest that 2
is a deacyl derivative of 1, in which Tig attached to C-4 of
Rha′ of 1 is cleaved. The sites of each ester linkage of the
acyl groups were confirmed using the HMBC spectrum of
2, with key cross-peaks observed between H-2 of Glc″ and
C-1 of Iva and methoxy protons and C-1 of Agl (Fig. 1).
Consequently, the structure of 2 was determined as methyl
3
4.48 dd (3.0, 9.0)
4.22 dd (9.0, 9.0)
4.30a)
4
5
6
1.64 d (6.5)
5.94 s
1.64 d (6.5)
5.90 s
Rha′-1
2
3
4
6.02 d (3.5), 5.99 d (3.5)
5.20a)
6.01 d (3.5), 6.02 d (3.5)
4.97a)
5.91 dd (10.0, 10.0),
5.91 dd (10.0, 10.0)
5.18a)
4.25 dd (9.0, 9.0)
5
6
5.00a)
1.75 d (6.5)
1.66 d (6.5)
2.75 dd (7.5, 14.5)
2.71 dd (5.0, 14.5)
4.44a)
3S,12S-dihydroxyhexadecanoate
12-O-β-D-glucopyranosyl-
Agl-2
2
2.75 dd (8.0, 15.0)
2.70 dd (4.0, 15.0)
4.43a)
(1→3)-O-(2-O-(3S,9R)-exogonoyl)-α-L-rhamnopyranosyl-
(1→2)-[O-(2-O-isovaleloyl)-β-D-glucopyranosyl-(1→3)]-O-β-D-
glucopyranosyl-(1→2)-[O-α-L-rhamnopyranosyl-(1→6)]-O-β-D-
glucopyranoside, as shown in Fig. 2.
3
12
3.86a)
3.87a)
16
0.90 t (7.5)
3.64 s
0.91 t (7.0)
OCH3
Exg-2
3.63 s
IOM-3 (3) was obtained as an amorphous powder and
exhibited an [M+Na]+ ion peak at m/z 1697, 84 mass units
(isovaleroyl residue) lager than that of 1, in the positive-
ion FAB-MS. The molecular formula of 3 was determined
as C78H130O38 based on the HR-positive-ion FAB-MS. The
1H-NMR spectrum of 3 was similar to that of 1, aside from
the appearance of signals because of one more isovaleroyl
3.12 dd (6.5, 16.0),
2.89 dd (6.5, 16.0)
2.87 dd (6.5, 16.0),
2.68 dd (6.5, 16.0)
4.71 dddd (6.5, 6.5, 6.5, 6.5), 4.63a)
4.30a), 4.05a)
1.32 d (7.0), 1.13 d (7.0)
7.43 q (7.0)
1.82 d (7.0)
2.02 s
3.19 dd (5.5, 15.5),
3.00 dd (5.5, 15.5)
2.90 dd (7.5, 15.5),
2.68 dd (7.5, 15.5)
4.74 m, 4.66 m
4.28a), 4.04a)
2
3
9
10
1.31 d (6.5), 1.14 d (6.5)
Tig-3
4
1
residue (Iva′). In the H-NMR spectrum of 3, compared with
that of 1, a remarkable downfield shift was observed for the
signal due to H-3 of the fourth glucosyl unit (Glc‴), whereas
the signals owing to H-2 and H-4 of Rha and H-2 of Glc″ of 3
were observed at chemical shifts similar to those of 1 (Table
3). Moreover, key cross-peaks between H-2 of Glc″ and C-1
of Iva, H-4 of Rha′ and C-1 of Tig, H-3 of Glc‴-3 and C-1 of
Iva′, and methoxy protons and C-1 of Agl were detected in the
HMBC spectrum of 3. These data indicate that 3 is a deriva-
tive of 1 with Iva′ attached to OH-3 of Glc‴ of 1.
5
Iva-2
2
2.72a)
2.72a)
2.54 dd (6.5, 16.0),
2.53 dd (6.5, 16.0)
2.33a)
2.51 m
3
4
5
2.33a)
1.19 d (6.5)
1.08 d (6.5)
1.18 d (7.0)
1.08 d (7.0)
δ in ppm from tetramethyl silane (TMS). Coupling constants (J) in Hz are given in
parentheses. a) Signals were overlapped with other signals.

Vol. 65, No. 1 (2017)
Chem. Pharm. Bull.
109
Table 2. 13C-NMR Spectral Data for 1–4 (in Pyridine-d5, 125MHz)
Position
1
2
3
4
Glc-1
102.6
78.7
102.5
79.1
102.7
78.7
102.7
79.2
2
3
79.9
79.6
80.0
80.1
4
71.8
72.0
71.8
71.7
5
76.5
76.3
76.6
76.6
6
68.2
68.2
68.3
68.2
Glc′-1
101.1
74.8, 74.9
86.9
101.2
75.5
87.1, 87.2
69.9
101.1
75.4
87.0
101.4
75.0
87.3
2
3
4
69.8
69.8
70.0
5
77.5
77.5
77.5
77.8
6
62.6
62.7
62.5
62.8
Glc″-1
100.3
74.5
100.5
74.6
100.4
74.6
100.5
74.6
2
3
4
75.3, 75.4
72.3
75.3
72.4
75.6
72.4
75.7
72.3
5
78.6
78.7
78.8
78.6
6
62.2
62.3
62.3
62.1
Glc‴-1
105.4
74.7
105.9
75.8
105.4, 105.3
72.8
105.9
74.7
2
3
78.2
78.3
78.6
78.1
4
5
71.8
78.2
71.7, 71.8
78.2
69.7
78.0
70.9
75.1
6
62.9
62.8
62.7
64.1
Rha-1
102.5
72.1
102.5
72.2
102.6
72.3
102.5
72.3
2
3
72.7
72.7
72.6
72.8
4
73.8
74.1
74.0
74.2
5
69.7
69.7
69.9
69.8
6
18.5
18.7
18.4
18.6
Rha′-1
96.1, 96.1
73.4, 73.2
74.9, 74.7
73.9
96.4, 96.6
73.3, 73.5
78.8
96.5, 96.4
73.1
96.5, 96.3
73.3, 73.1
74.7
2
3
4
75.4
74.2
73.3
74.0
5
6
67.2
18.3
69.3
18.8
67.2
18.7
67.3
18.5
Agl-1
2
172.8
43.3
172.9
43.4
172.9
43.4
172.9
43.4
3
68.2
68.3
68.3
68.3
12
81.6
81.3
81.6
81.5
16
14.3
14.4
14.3
14.4
OCH3
51.2
51.3
51.3
51.3
Exg-1 171.3, 171.0
171.4, 171.0
43.2, 41.4
75.8, 74.3
115.2, 115.1
76.1, 74.2
23.4, 21.3
171.4, 171.1
43.2, 41.2
75.6, 73.8
115.2, 115.1
76.1, 74.2
23.4, 21.2
167.6
129.1
138.5
14.5
170.6, 170.4
43.3, 41.1
75.5, 73.8
115.1, 115.0
76.0, 74.2
23.3, 21.3
167.6
129.2
138.1
14.6
2
3
6
9
43.2, 41.0
75.5, 74.2
115.2, 115.0
76.0, 74.1
23.3, 21.2
167.5
10
Tig-1
2
129.1
Fig. 1. 1H–13C Long-Range Correlations Observed in the HMBC Spec-
tra of 1–3 (in Pyridine-d5, 500MHz)
3
138.2
4
14.5
5
12.8
172.1
43.0
25.5
22.6
22.4
12.7
172.1
43.1
25.3
22.6
22.5
172.6
43.9
25.6
22.5
12.9
172.1
43.1
25.6
22.6
22.5
173.1
43.3
25.9
22.6
Partial deacylation of 3 was performed in order to de-
termine the sites of ester linkages of the respective or-
ganic acids. Compound 3 was refluxed with 3% trieth-
ylamine in MeOH for 2h, and the reaction mixture was
separated by HPLC, affording 1. Consequently, the structure
of 3 was identified as methyl 3S,12S-dihydroxyhexadecanoate
12-O-(3-O-isovaleloyl)-β-D-glucopyranosyl-(1→3)-O-(2-O-
(3S,9R)-exogonoyl,4-O-tigloyl)-α-L-rhamnopyranosyl-(1→2)-
[O-(2-O-isovaleroyl)-β-D-glucopyranosyl-(1→3)]-O-β-D-
glucopyranosyl-(1→2)-[O-α-L-rhamnopyranosyl-(1→6)]-O-β-D-
glucopyranoside, as shown in Fig. 2.
Iva-1
172.1
43.4
25.4
22.7
22.5
2
3
4
5
Iva′-1
2
3
4
5
22.5
22.6
δ in ppm from TMS.
IOM-4 (4) was obtained as an amorphous powder. The HR-

110
Chem. Pharm. Bull.
Vol. 65, No. 1 (2017)
Table 3. 1H-NMR Spectral Data for 3 and 4 (in Pyridine-d5, 500MHz)
Position
3
4
Glc-1
4.81 d (8.0)
4.29a)
4.85 d (7.5)
4.32a)
2
3
4
5
4.43a)
4.42a)
3.87a)
3.97a)
3.91 dd (9.0, 9.0)
4.00a)
6
6
4.61 brd (11.0)
4.06 dd (7.0, 11.0)
5.74 d (8.0)
4.12 dd (8.0, 9.0)
3.96 dd (9.0, 9.0)
3.84 dd (9.0, 9.0)
3.68 m
4.62 brd (10.5)
4.08 dd (6.5,10.5)
5.80 d (7.5)
4.18 dd (7.5, 8.5)
4.12a)
Glc′-1
2
3
4
5
3.89a)
3.79 m
4.36 dd (2.5, 12.0)
4.16a)
6
4.31a)
6
Glc″-1
2
4.13a)
5.14 d (7.5)
5.53 dd (7.5, 9.0)
5.18 d (8.0)
5.55 dd (8.0, 10.0),
5.54 dd (8.0, 10.0)
4.42a)
3
4
5
6
4.41a)
4.12a)
4.12a)
4.13a)
Fig. 2. Structures of 1–4
4.13a)
4.53 d (11.0)
4.27a)
4.50a)
positive-ion FAB-MS showed that its molecular formula was
6
4.28a)
the same as 3. The H- and 13C-NMR spectra of 4 were simi-
1
Glc‴-1
5.16 d (7.5)
3.87a)
5.12 d (7.5)
3.88a)
lar to those of 3 and showed signals corresponding to the pres-
ence of one each of Tig, Exg, and methoxy group and two iso-
valeroyl residues (Tables 2, 3). From these data, 4 was deduced
to be a positional isomer of 3 concerning ester linkage. The
1H-NMR spectrum of 4 exhibited, in comparison with that of
3, downfield shifts for the signals assignable to H2-6 of Glc‴
along with an upfield shift for the signal assignable to H-3 of
Glc‴. In contrast, the signals of H-2 and H-4 of Rha and H-2
of Glc″ appeared at positions similar to those in 3. Further,
the treatment of 4 with mild alkali in a similar manner as
for 3 afforded 1. Therefore, 4 was a positional isomer of 3, in
which the isovaleroyl residue of 4 was at the OH-6 of Glc‴
rather than at the OH-3 of Glc‴. Consequently, the structure
of 4 was assigned as methyl 3S,12S-dihydroxyhexadecanoate
12-O-(6-O-isovaleloyl)-β-D-glucopyranosyl-(1→3)-O-(2-O-
(3S,9R)-exogonoyl,4-O-tigloyl)-α-L-rhamnopyranosyl-(1→2)-
[O-(2-O-isovaleroyl)-β-D-glucopyranosyl-(1→3)]-O-β-D-
glucopyranosyl-(1→2)-[O-α-L-rhamnopyranosyl-(1→6)]-O-β-D-
glucopyranoside, as shown in Fig. 2.
The convolvulin from I. purga was speculated to be an
oligomer of an acylated glycosidic acid.14) Recently, jalpinoside,
which was separated as a minor constituent from the convol-
vulin fraction of I. purga, was reported a macrocyclic bisdes-
moside resin glycoside.15) However, jalapinoside is considered
as an acylated glycosidic acid methyl ester monomer. The
1H-NMR spectrum of jalapinoside in the Supporting Informa-
tion indicates signal due to a methoxy group, which shows a
correlation with a carbonyl carbon (C-1) of aglycone moiety
in its HMBC spectrum.15) IOM-1–IOM-4 were all regarded
as the methyl ester monomers of the acylated glycosidic acid.
Further, negative-ion FAB-MS and HR-negative-ion electro-
spray ionization-time-of-flight (ESI-TOF)-MS of the crude
convolvulin fraction exhibited intense ion peaks at m/z 1659
and 1659.8010 (calcd for C77H127O38–, 1659.8011), correspond-
ing to the values of an [M–H]– ion peak of the demethyl de-
rivatives of 3 and 4, respectively. In addition, a distinct peak
wasn’t detected in the region of about m/z 1700 to 4000 in
the negative-ion FAB-MS of the crude convolvulin fraction.
Therefore, a part of the crude convolvulin fraction might be a
mixture of monomers composed of free carboxylic acid forms
2
3
4
5
6
6
5.60 dd (9.0, 9.0)
3.98a)
4.01a)
3.98a)
4.00a)
4.88 dd (2.0, 11.5)
4.74a)
3.98a)
4.46a)
4.11a)
Rha-1
5.44 s
4.54 s
5.44 s
2
3
4
5
4.54 d (3.0)
4.48 dd (3.0, 9.0)
4.25 dd (9.0, 9.0)
4.30a)
1.64 d (6.5)
5.99a)
5.99a)
5.12a)
5.89 dd (10.0, 10.0),
5.88 dd (10.0, 10.0)
5.16a)
1.65 d (7.0)
2.75 dd (8.0, 15.0)
2.71 dd (4.5, 15.0)
4.44a)
4.48 dd (3.0, 9.0)
4.25 dd (9.0, 9.0)
4.31a)
1.66 d (6.0)
5.94 s
6
Rha′-1
2
3
4
6.07 d (3.0), 6.05 d (3.0)
5.11a)
5.88 dd (9.5, 9.5),
5.88 dd (9.5, 9.5)
5.20 m
5
6
1.66 d (6.0)
2.75 dd (7.5, 14.5)
2.71a)
Agl-2
2
3
12
16
OCH3
Exg-2
4.43a)
3.87a)
3.88a)
0.91 t (7.5)
3.63 s
0.91 t (7.0)
3.63 s
3.12 dd (6.5, 16.0),
2.90 dd (6.5, 16.0)
2.88a), 2.69a)
3.16 dd (4.5, 16.0),
2.93 dd (5.0, 16.0)
2.90 dd (9.0, 16.0),
2.67 dd (8.5, 16.0)
4.72 m, 4.70 m
4.25a), 4.02a)
1.27 d (6.5), 1.15 d (6.5)
7.42 q (7.0)
1.82 d (7.0)
2.05 brs
2
3
4.73 dddd (6.5, 6.5, 6.5, 6.5), 4.65a)
4.27a), 4.01a)
1.33 d (6.5), 1.14 d (6.5)
7.39 q (7.0)
1.81 d (7.0)
1.98 s
9
10
Tig-3
4
5
Iva-2
2
2.70a)
2.53 dd (6.5, 16.0),
2.52 dd (6.5, 16.0)
2.35a)
2.74a)
2.53a)
3
4
5
2.35a)
1.19 d (7.0)
1.08 d (7.0)
2.20a)
2.20a)
2.11a)
1.19 d (7.0)
1.08 d (7.0)
2.33 d (7.5)
2.33 d (7.5)
2.10 m
Iva-2
2
3
4
5
0.89 d (6.5)
0.89 d (6.5)
0.94 d (6.5)
0.94 d (6.5)
δ in ppm from TMS. Coupling constants (J) in Hz are given in parentheses. a)
Signals were overlapped with other signals.

Vol. 65, No. 1 (2017)
Chem. Pharm. Bull.
111
corresponding to 3 and 4, etc.
12.1 (tiglic acid) for 1]. A part of the organic acid fraction
was methylated with diazomethane-ether and then the reaction
mixture was analyzed by GC [column, OV-1 (1%), GL Scienc-
Experimental
The instruments and materials used were as cited in the es Inc., Tokyo, Japan, 3.2mm i.d.×2m glass column; column
preceding report16) unless otherwise specified. ESI-TOF-MS temperature, 130°C; carrier gas, N2 1.5 kg/cm2; tR (min): 7.6
data was collected using a Q-Exactive Plus (Thermo Fisher (methyl exigonate) for 1 and 2].9)
Scientific K.K., Kanagawa, Japan) mass spectrometer.
Partial Deacylation of 3 and 4 Solutions of 3 (4mg) and
Treatment of the Convolvulin Fraction with Indium(III) 4 (3mg) in 3% triethylamine–MeOH (1mL) were refluxed for
Chloride in MeOH and Separation of 1–4 Indium(III) 2h, respectively. After removal of the solvent, reaction mix-
chloride (2.0g) was added to a solution of the crude covolvu- tures were each subjected to HPLC (column, Inertsil ODS-3,
lin fraction9) (4.00g) from the roots of I. operculata in MeOH GL Sciences Inc., 4.6mm i.d.×250mm) using 85% MeOH to
(160mL), and the mixture was refluxed for 25d, while being give 1 (2mg from 3, 1mg from 4), which was identified by
1
monitored by TLC. The solvent was removed under reduced comparing of its H-NMR spectrum with that of an authentic
pressure, and the residue was subjected to Sephadex LH-20 sample.
colum chromatography (CC) eluted with MeOH to give frac-
HPLC Analyses of 1a and b Each HPLC analysis
tions 1 (1189mg), 2 (2450mg), and 3 (19mg). CC of fraction (column, COSMOSIL π-nap, Nacalai Tesque, Inc., 4.6mm
2 on silica gel eluted with a gradient of mixtures of CHCl3– i.d.×250mm; solvent, 80% MeOH; flow rate 0.8mL/min) of
MeOH–H2O (10:2:0.1, 8:2:0.2, 7:3:0.5, 6:4:1, 0:1:0) 1a and b afforded two peaks in which the Rt values (27.6min
afforded 1 (56mg) and fractions 2-1–2-63. Fractions 2-16 and 30.3min) matched with those of 1a and b, respectively.
(427mg) and 2-43 (89mg) were each subjected to HPLC [col-
umn, COSMOSIL 5C18-AR-II (Nacalai Tesque, Inc., Kyoto,
Acknowledgments We express our appreciation to Mr. H.
Japan, 20mm i.d.×250mm)] using 85% MeOH for fractions Harazono of Fukuoka University for the measurement of the
2-16 and 80% MeOH for fraction 2-43 as eluent to yield 4 FAB-MS and to Ms. Mihoko Yamaguchi of Thermo Fisher
(49mg) and 3 (57mg) from fraction 2-16 and 2 (13mg) from Scientific, Inc. for the measurement of the ESI-TOF-MS. This
fraction 2-43. Compounds 1 (25mg), 2 (3mg), 3 (25mg), and research was supported in part by a Grant-in-Aid for Scientific
4 (15mg) were each subjected to HPLC [column, COSMOSIL Research (C) (No. 16K08306) from Japan Society for the Pro-
π-nap (Nacalai Tesque, Inc., 20mm i.d.×250mm)] using 75% motion of Science.
MeOH for 2, 80% MeOH for 1 and 4, and 85% MeOH for 3
as eluent to yield 1a (4mg) and 1b (7mg) from 1, 2a (1mg)
Conflict of Interest The authors declare no conflict of
and 2b (1mg) from 2, 3a (8mg) and 3b (5mg) from 3, and 4a interest.
(4mg) and 4b (5mg) from 4.
1
References
The H-NMR spectra of 1a, 2a, 3a, and 4a were superim-
posable on those of 1b, 2b, 3b, and 4b, respectively.
IOM-1 (1)
1) Mayer W., Justus Liebigs Ann. Chem., 95, 129–176 (1855).
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Amorphous powder. [α]2D6 –19.1° (c=1.0, MeOH). Positive-
ion FAB-MS m/z: 1613 [M+Na]+. HR-positive-ion FAB-MS
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m/z: 1613.7570 (Calcd for C73H122O37Na+, 1613.7557). H-NMR
1
spectral data: see Table 1. 13C-NMR spectral data: see Table 2.
IOM-2 (2)
Amorphous powder. [α]1D9 –20.6° (c=1.7, MeOH). Positive-
ion FAB-MS m/z: 1531 [M+Na]+. HR-positive-ion FAB-MS
m/z: 1531.7135 (Calcd for C68H116O36Na+, 1531.7139). H-NMR
1
spectral data: see Table 1. 13C-NMR spectral data: see Table 2.
IOM-3 (3)
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Amorphous powder. [α]2D6 –21.3° (c=1.7, MeOH). Positive-
ion FAB-MS m/z: 1697 [M+Na]+. HR-positive-ion FAB-MS
m/z: 1697.8157 (Calcd for C78H130O38Na+, 1697.8132). H-NMR
1
spectral data: see Table 3. 13C-NMR spectral data: see Table 2.
IOM-4 (4)
Amorphous powder. [α]2D6 –2.2° (c=1.5, MeOH). Positive-
ion FAB-MS m/z: 1697 [M+Na]+. HR-positive-ion FAB-MS
m/z: 1697.8157 (Calcd for C78H130O38Na+, 1697.8132). H-NMR
1
spectral data: see Table 3. 13C-NMR spectral data: see Table 2.
Alkaline Hydrolysis of 1 and 2 The alkaline hydrolysis
was applied in a slightly modified manner previously re-
ported.9) Briefly, alkaline hydrolysis (95°C, 1h) of 1 (15mg)
and 2 (10mg) with 3% KOH furnished organic acid fraction
and glycosidic acid (10mg from 1, 5mg from 2), respectively.
Both glycosidic acids derived from 1 and 2 were identified as
1
5 based on their H-NMR spectra.9) The organic fraction was
analyzed by GC9) [tR (min): 5.2 (isovaleric acid) for 1 and 2,