Three new aromatic glycosides from the ripe fruit of cherry tomato

Article abstract of DOI:10.1007/s11418-010-0436-3

Three new aromatic glycosides were isolated from the ripe fruit of cherry tomato [Lycopersicon esculentum var. cerasiforme (Dunal) Alef. (Solanaceae)] along with six known aromatic glycosides and one known steroidal alkaloid glycoside. Their chemical structures were determined on the basis of spectroscopic data as well as chemical evidence.

Full text of DOI:10.1007/s11418-010-0436-3

J Nat Med (2010) 64:500–505
DOI 10.1007/s11418-010-0436-3
NOTE
Three new aromatic glycosides from the ripe fruit
of cherry tomato
Masateru Ono
Shin Yasuda
Hitoshi Yoshimitsu
•
Yuki Shiono
Tsuyoshi Ikeda
Toshihiro Nohara
•
Takayuki Tanaka
•
•
Chikako Masuoka
Junei Kinjo
•
•
Masafumi Okawa
•
•
•
Received: 8 April 2010 / Accepted: 25 May 2010 / Published online: 24 June 2010
Ó The Japanese Society of Pharmacognosy and Springer 2010
Abstract Three new aromatic glycosides were isolated
from the ripe fruit of cherry tomato [Lycopersicon escu-
lentum var. cerasiforme (DUNAL) ALEF. (Solanaceae)] along
with six known aromatic glycosides and one known ste-
roidal alkaloid glycoside. Their chemical structures were
determined on the basis of spectroscopic data as well as
chemical evidence.
and in cooking, and the latter as a fresh vegetable. We earlier
reported the isolation and structural elucidation of a steroidal
alkaloid glycoside, esculeoside A, from the ripe pink color-
type tomato (L. esculentum, ‘Momotaro’); a solanocapsine-
type glycoside, esculeoside B, from the ripe red color-type
tomato (L. esculentum, ‘Italian San Marzano’); six steroidal
glycosides, esculeosides A, B, C, and D, lycoperoside G,
and 3-O-b-lycotetraosyl 3b,26-dihydroxy-cholestan-16,
22-dione 26-O-a-L-arabinopyranosyl-(1 ? 6)-b-D-gluco-
pyranoside, and six aromatic compounds, zizybeoside I,
benzoyl alcohol b-gentiobioside, rutin, methyl caffeate,
phenylalanine, and 4-hydroxyphenyl b-D-glucopyranosyl-
(1 ? 2)-b-D-glucopyranoside, from the ripe cherry tomato
Keywords Lycopersicon esculentum ꢀ Cherry tomato ꢀ
Solanaceae ꢀ Aromatic glycoside ꢀ Methyl salicylate
Introduction
[
L. esculentum var. cerasiforme (DUNAL) ALEF.]; and two
Tomato, the fruit of Lycopersicon esculentum (syn. Solanum
lycopersicum L., Solanaceae), is most widely used as a fresh
vegetable and for cooking. The species of tomato on the
market are roughly classified into two groups, red color-type
and pink color-type; the former is mainlyused for pasta sauce
pregnane glycosides, tomato pregnane and 3-O-b-lycotet-
raosyl 5a-pregna-3b,16b-diol-20-one, from the overripe
cherry tomato [1–6]. As for the constituents of the tomato,
differences were observed among cultivars as was men-
tioned above. Therefore, as part of our continuing study on
the chemical constituents of the tomato, we examined the
constituents of the ripe fruit of cherry tomato (‘Komomo’).
The present paper deals with the isolation and structural
characterization of three new aromatic glycosides along with
seven known compounds from the ripe fruit.
M. Ono (&) ꢀ Y. Shiono ꢀ T. Tanaka ꢀ C. Masuoka ꢀ S. Yasuda
School of Agriculture, Tokai University,
5435 Minamiaso, Aso, Kumamoto 869-1404, Japan
e-mail: mono@agri.u-tokai.ac.jp
T. Ikeda
Faculty of Medical and Pharmaceutical Sciences,
Kumamoto University, 5-1 Oe-honmachi,
Kumamoto 862-0973, Japan
Results and discussion
The ripe fruit of cherry tomato was smashed with H O in a
2
mixer then filtered through filter paper to give a filtrate,
which was subjected to Diaion HP20, Sephadex LH-20,
M. Okawa ꢀ J. Kinjo
Faculty of Pharmaceutical Sciences, Fukuoka University,
8
-19-1 Nanakuma, Jonan-ku, Fukuoka 814-0180, Japan
Chromatorex NH , Chromatorex ODS, and silica gel col-
2
umn chromatography, as well as HPLC on ODS to afford
nine aromatic glycosides (1–9) and one steroidal alkaloid
glycoside (10).
H. Yoshimitsu ꢀ T. Nohara
Faculty of Pharmaceutical Sciences, Sojo University,
4
-22-2 Ikeda, Kumamoto 860-0082, Japan
1
23

J Nat Med (2010) 64:500–505
501
Compound 1 was obtained as an amorphous powder and
due to one phenyl group [d 139.5, 129.6 (92), 128.6 (92),
126.4], three anomeric carbons (d 106.4, 105.5, 102.9), and
one methylene carbon (d 36.6). In the same manner as for
1, these H- and C-NMR signals were examined in detail
and the data suggested that 2 was composed of 1 mol of
phenethyl alcohol, 2 mol of glucose, and 1 mol of arabi-
?
exhibited an [M ? Na] ion peak at m/z 631 in the positive
FAB–MS; the high-resolution (HR) positive FAB–MS
1
13
indicated the molecular formula of 1 to be C H O . The
2
5 36 5
1
H-NMR spectrum of 1 revealed the presence of four
aromatic protons [d 7.84 (1H, d, J = 1.5, 7.5 Hz), 7.67
1H, d, J = 8.0 Hz), 7.46 (1H, ddd, J = 1.5, 7.5, 8.0 Hz),
.88 (1H, dd, J = 7.5, 7.5 Hz)] assigned to an ortho-
substituted phenyl group, three anomeric protons [d 5.62
1H, d, J = 7.5 Hz), 5.56 (1H, d, J = 8.0 Hz), 4.90 (1H, d,
J = 7.5 Hz)], and one methoxy group [d 3.90 (3H, s)]. The
C-NMR spectrum of 1 gave 25 carbon signals, including
those corresponding to one carboxyl carbon (d 166.3), six
aromatic carbons (d 157.4, 134.1, 131.6, 121.5, 121.0,
(
nose. The data also indicated that the glycosidic linkages of
4
6
the glucose units were b in C conformations and that of
1
4
the arabinose unit was a in a C conformation. Glyco-
1
1
3
(
sylation shifts [7, 8] in the C-NMR data of 2 were
observed at C-2 (?9.0 ppm) and C-6 (?6.9 ppm) of Glc.
In addition, the HMBC spectrum of 2 showed the key
1
3
0
correlations between H-1 of Glc and C-2 of Glc; H-1 of
arabinosyl unit (Ara) and C-6 of Glc; and H-1 of Glc and
C-8 of Ag. Consequently, 2 was elucidated as phenethyl
alcohol 8-O-b-D-glucopyranosyl-(1 ? 2)-[O-a-L-arabino-
pyranosyl-(1 ? 6)]-O-b-D-glucopyranoside.
Compound 3 was obtained as an amorphous powder.
?
The positive FAB–MS of 3 showed an [M ? Na] ion
1
16.4), three anomeric carbons (d 105.9, 105.7, 100.0), and
1
13
one methoxy carbon (d 52.0). These H- and C-NMR
1
1
signals were assigned with the aid of H– H COSY,
HMQC, and HMBC spectra (Tables 1, 2), and 1 was
determined to be a methyl salicylate 2-O-triglycoside,
which was composed of 1 mol of pentose and 2 mol of
hexose.
peak at m/z 587, which was 14 mass units (CH ) smaller
2
than that of 2, and the molecular formula of 3 was deter-
mined to be C H O using HR-positive FAB–MS. The
Acidic hydrolysis of 1 afforded D-xylose and D-glucose,
which were confirmed by optical rotation using chiral
detection in HPLC analyses. The coupling constants of
signals due to anomeric and methine protons of xylosyl and
2
4 36 15
1
13
H- and C-NMR spectra of 3 were superimposable on
those of 2, except for the appearance of the signals due to a
benzyl alcohol unit and the lack of the signals due to a
phenethyl alcohol unit. On the basis of these data, 3 was
concluded to be benzyl alcohol 7-O-b-D-glucopyranosyl-
(1 ? 2)-[O-a-L-arabinopyranosyl-(1 ? 6)]-b-D-glucopyran-
oside.
1
glucosyl units in the H-NMR spectrum indicated that all
the monosaccharide units were of the pyranose type, and
4
all glycosidic linkages were b in C conformations. The
1
1
3
C-NMR data for the monosaccharide units were com-
pared with those of corresponding methyl pyranosides in
the literature [7]. Glycosylation shifts [7, 8] were observed
at C-2 (?7.9 ppm) and C-6 (?7.0 ppm) of the inner glu-
cosyl unit (Glc). Moreover, key correlations were observed
Compounds 4–10 were identified as kaempferol
3-O-rutinoside (4) [9], rutin (5) [9], quercetin 3-O-b-apio-
furanosyl-(1 ? 2)-O-[a-rhamnopyranosyl-(1 ? 6)]-b-gluco-
pyranoside (6) [9], phenethyl alcohol b-gentiobioside (7)
[10], zizybeoside I (8) [11], dihydroconiferyl 4-O-b-D-
glucopyranoside (9) [12], and esculeoside B (10) [2],
respectively, based on comparison of their physical and
spectral data with authentic samples or those already
reported.
0
between H-1 of the terminal glucosyl unit (Glc ) and C-2 of
Glc; H-1 of xylosyl unit (Xyl) and C-6 of Glc; and H-1 of
Glc and C-2 of aglycone moiety (Ag) in the HMBC
spectrum of 1 (Fig. 1). The structure of 1 was therefore
defined as methyl salicylate 2-O-b-D-glucopyranosyl-
(1 ? 2)-[O-b-D-xylopyranosyl-(1 ? 6)]-O-b-D-glucopyra-
noside (Fig. 2).
To gain insight into the biological functions of tomato
constituents, the antioxidative properties of MeOH-eluted
Compound 2 was obtained as an amorphous powder,
and gave D-glucose and L-arabinose on acidic hydrolysis.
fractions (using a Diaion HP20 column) obtained from H O
2
extracts of ‘Komomo’ and ‘Momotarofight’ (L. esculentum)
cultivars were analyzed. The ‘Komomo’ fraction revealed
ca. threefold lower 1,1-diphenyl-2-picrylhydrazyl (DPPH)
radical scavenging activity (EC50 value of 48.5 lg/ml)
than that of the ‘Momotarofight’ fraction (EC50 value of
16.5 lg/ml) (Fig. 3a). In a superoxide anion radical scav-
enging assay, the activity of the ‘Komomo’ fraction was ca.
1.4-fold higher (EC50 value of 174 lg/ml) than that of the
‘Momotarofight’ fraction (EC50 value of 246 lg/ml)
(Fig. 3b). It is conceivable that the antioxidative activities
of these tomato fractions may be because of the different
compositions and/or amount of active compounds present in
?
The positive FAB–MS of 2 showed an [M ? Na] ion
peak at m/z 601. The molecular formula of 2 was deter-
mined to be C H O using HR-positive FAB–MS. The
2
5 38 15
1
H-NMR spectrum of 2 indicated signals due to five aro-
matic protons [d 7.32 (2H, d, J = 8.5 Hz), 7.26 (2H, dd,
J = 8.5, 8.5 Hz), 7.15 (1H, t, J = 8.5 Hz)], one oxygen-
ated methylene group [d ca. 4.27 (1H), 3.78 (1H, ddd,
J = 7.5, 8.5, 8.5 Hz)], one methylene group [d 3.09 (2H,
m)], and three anomeric protons [d 5.29 (1H, d,
J = 8.0 Hz), 4.86 (1H, d, J = 7.0 Hz), 4.82 (1H, d,
1
3
J = 8.0 Hz)]. The C-NMR spectrum of 2 showed signals
123

5
02
J Nat Med (2010) 64:500–505
1
Table 1 H-NMR spectral data
for 1–3 (in pyridine-d
00 MHz)
1
2
3
5
,
5
Ag-2
Ag-3
Ag-4
Ag-5
Ag-6
Ag-7
Ag-7
Ag-8
Ag-8
OCH
7.32 d (8.5)
7.26 dd (8.5, 8.5)
7.15 t (8.5)
7.72 d (7.5)
7.67 d (8.0)
7.38 dd (7.5, 7.5)
7.22 t (7.5)
7.46 ddd (1.5, 7.5, 8.0)
6.88 dd (7.5, 7.5)
7.84 dd (1.5, 7.5)
7.26 dd (8.5, 8.5)
7.32 d (8.5)
3.09 m
7.38 dd (7.5, 7.5)
7.72 d (7.5)
5.18 d (12.0)
4.81 d (12.0)
3.09 m
ca. 4.27
3.78 ddd (7.0, 8.5, 8.5)
3
3.90 s
Glc-1
Glc-2
Glc-3
Glc-4
Glc-5
Glc-6
Glc-6
5.62 d (7.5)
4.82 d (8.0)
4.06 dd (8.0, 9.5)
ca. 4.25
4.94 d (7.5)
4.47 dd (7.5, 9.0)
4.35 dd (9.0, 9.0)
4.14 dd (9.0, 9.0)
ca. 4.21
4.15 dd (7.5, 9.0)
4.21 dd (9.0, 9.0)
4.12 dd (9.0, 9.0)
4.00 ddd (2.5, 6.0, 9.0)
4.80 dd (2.5, 11.0)
4.26 dd (6.0, 11.0)
5.31 d (7.5)
ca. 4.10
3.96 ddd (2.0, 5.5, 9.5)
4.77 dd (2.0, 11.0)
ca. 4.23
4.75 dd (1.5, 11.5)
ca. 4.25
0
Glc -1
0
5.56 d (8.0)
5.29 d (8.0)
ca. 4.10
Glc -2
0
4.08 dd (8.0, 9.0)
ca. 4.26
4.10 dd (7.5, 9.0)
4.26 dd (9.0, 9.0)
ca. 4.30
Glc -3
0
ca. 4.25
Glc -4
0
4.30 dd (8.0, 8.0)
3.99 m
ca. 4.29
Glc -5
0
ca. 3.95
3.91 ddd (3.0, 4.5, 9.0)
4.48 dd (3.0, 11.5)
4.41 dd (4.5, 11.5)
Glc -6
0
4.34 dd (4.5, 11.5)
ca. 4.24
4.53 dd (2.5, 11.5)
4.39 dd (5.0, 11.5)
Glc -6
Xyl-1
Xyl-2
Xyl-3
Xyl-4
Xyl-5
Xyl-5
Ara-1
Ara-2
Ara-3
Ara-4
Ara-5
Ara-5
4.90 d (7.5)
4.00 dd (7.5, 9.0)
4.08 dd (9.0, 9.0)
ca. 4.19
ca. 4.25
3.56 dd (11.0, 11.0)
4.86 d (7.0)
4.91 d (7.0)
d in ppm from TMS (coupling
constants (J) in Hz are given in
parentheses)
4.44 dd (7.0, 8.5)
4.12 dd (3.0, 8.5)
ca. 4.29
4.48 dd (7.0, 8.5)
4.15 dd (3.5, 8.5)
ca. 4.31
Glc glucopyranosyl, Xyl
xylopyranosyl, Ara
arabinopyranosyl, Ag aglycone
moiety
ca. 4.28
ca. 4.30
3.72 dd (3.0, 13.0)
3.74 dd (2.5, 13.0)
1
3
the fruits of both cultivars. Investigations into the biological
functions of the isolated compounds from the fruit of
C-NMR spectra were recorded with JEOL ECA 500
spectrometer (JEOL), and chemical shifts are given on a d
(ppm) scale with tetramethylsilane (TMS) as an internal
standard. Silica gel 60 (Merck, Art. 9385; Merck,
Darmstadt, Germany), Sephadex LH-20 (Pharmacia Fine
Chemicals, Uppsala, Sweden), Chromatorex ODS (Fuji
Silysia Chemical, Ltd., Aichi), Chromatorex NH2 (Fuji
Silysia Chemical, Ltd.), and Diaion HP20 (Mitsubishi
Chemical Industries Co., Ltd., Tokyo) were used for
column chromatography. HPLC separation was run on
a Shimadzu LC-10AS micro pump (Shimadzu, Kyoto)
with Shimadzu RID-10A RI-detector (Shimadzu). For
HPLC column chromatography, COSMOSIL 5C18-AR-II
‘Komomo’ and constituents from the fruit of ‘Momotarofight’
are now in progress.
Experimental
General
Optical rotations were performed with a JASCO DIP-
1
000 KYU digital polarimeter (JASCO, Tokyo). MS were
1
recorded on a JEOL JMS-700T (JEOL, Tokyo). H- and
1
23

J Nat Med (2010) 64:500–505
503
13
Table 2 C-NMR spectral data for 1–3 (in pyridine-d , 125 MHz)
5
1
2
3
1
2
3
0
Ag-1
Ag-2
Ag-3
Ag-4
Ag-5
Ag-6
Ag-7
Ag-8
OCH
121.0 139.5 138.9 Glc -1 105.7 106.4 106.6
0
157.4 129.6 128.1 Glc -2
0
76.8
78.1
71.2
78.3
62.3
105.9
75.0
78.1
70.8
67.1
76.7
78.7
71.4
78.2
62.8
76.8
78.6
71.4
78.1
62.6
116.4 128.6 128.6 Glc -3
0
134.1 126.4 127.6 Glc -4
0
121.5 128.6 128.6 Glc -5
0
131.6 129.6 128.1 Glc -6
166.3
52.0
36.6
70.8
70.9 Xyl-1
Xyl-2
3
Xyl-3
Glc-1
Glc-2
Glc-3
Glc-4
Glc-5
Glc-6
100.0 102.9 102.2 Xyl-4
82.7
77.8
71.1
77.5
69.5
83.8
77.9
71.6
76.8
69.4
84.1 Xyl-5
78.0 Ara-1
71.5 Ara-2
76.9 Ara-3
69.5 Ara-4
Ara-5
105.5 105.7
72.3
74.4
69.2
66.6
72.4
74.4
69.3
66.7
d in ppm from TMS
Glc glucopyranosyl, Xyl xylopyranosyl, Ara arabinopyranosyl, Ag
aglycone moiety
1
13
Fig. 1 H– C long-range
correlations observed for 1 in
the HMBC spectrum (in
HO
HO
O
O
O
HO
HO
O
O
HO
O
OCH3
pyridine-d
5
, 500 MHz)
O
OH
HO
Fig. 3 Antioxidative activities of two different tomato fractions from
‘Komomo’ and ‘Momotarofight’ cultivars on the DPPH radical (a)
and superoxide anion radical scavenging analyses (b)
HO OH
HMBC: H
C
paper to give a filtrate, which was subjected to Diaion
5
4
3
6
1
HO
2
6
3
HP20 column chromatography (H O, MeOH, acetone) to
2
Xyl
Ara
7
1
4
HO
HO
O
O
O
O
O
Glc
O
2
Glc
O
2: R = O
8
HO
afford the MeOH-eluted fraction (fr.) and acetone-eluted fr.
The MeOH-eluted fr. (17.9 g) was chromatographed over a
Sephadex LH-20 column (90% MeOH, MeOH) to give frs.
HO
HO
HO
HO
R
5
HO
O
OCH3
HO
2
3
5
O
O
7
1
4
Glc'
O
1
Glc'
O
3: R = O
6
OH
OH
HO
HO OH
HO
HO OH
1
–4. Chromatography of fr. 1 (16.8 g) over a Sephadex
LH-20 column (30% MeOH, MeOH) furnished frs. 1.1–
.4. Fraction 1.3 (1.1 g) was chromatographed over a
Chromatorex NH column [CHCl –MeOH–H O (8:2:0.1,
Fig. 2 Structures of 1–3
1
2
3
2
(
Nacalai Tesque Inc., Kyoto, 20-mm i.d. 9 250 mm) was
7:3:0.5, 6:4:1, 5:5:1, 3:6:1), 100% MeOH, 90% MeOH,
80% MeOH, 70% MeOH, 60% MeOH, 50% MeOH, 40%
used.
MeOH, 30% MeOH, 20% MeOH, H O] to give frs. 1.3.1–
2
Plant material
1.3.19. Chromatography of fr. 1.4 (9.1 g) over a Chro-
matorex NH column under the same conditions to those of
2
The ripe fruits of cherry tomato (‘Komomo’) were col-
lected in October 2008 at the farm of Tokai University,
Kumamoto prefecture, Japan.
fr. 1.3 furnished frs. 1.4.1–1.4.20. Fractions 1.3.5
(130 mg), 1.3.8 (419 mg), 1.4.5 (268 mg), 1.4.7 (356 mg),
and 1.4.11 (258 mg) were each subjected to HPLC (30%
MeOH) to afford 7 (7 mg) and 8 (3 mg) from fr. 1.3.5, 10
(32 mg) from fr. 1.3.8, 9 (7 mg) from fr. 1.4.5, 7 (17 mg)
from fr. 1.4.7, and 1 (6 mg), 2 (8 mg), and 3 (3 mg) from
fr. 1.4.11. Fraction 2 (0.8 g) was chromatographed over a
Chromatorex ODS column (40% MeOH, 50% MeOH, 60%
Extraction and isolation
The ripe fruits (17836 g) of cherry tomato were smashed
with H O (26650 ml) in a mixer then filtered through filter
2
123

5
04
J Nat Med (2010) 64:500–505
MeOH, 70% MeOH, 80% MeOH, 90% MeOH, 100%
MeOH) to furnish frs. 2.1–2.11 and 5 (11 mg). Chroma-
tography of fr. 2.6 (89 mg) over a Diaion HP20 column
on the following method [13]. Briefly, the reaction was
started by the addition of DPPH into the assay mixture
containing varying concentrations of test samples, then
allowed to proceed for 30 min at room temperature. The
absorbance of the resulting solution was measured at
(
(
H O, MeOH, acetone) furnished 6 (9 mg), fr. 2.6.1
2
58 mg), and fr. 2.6.2 (11 mg). Fraction 2.10 (49 mg) was
-
subjected to silica gel column chromatography [CHCl3–
517 nm. The superoxide anion (O2 ) radical scavenging
MeOH–H O (10:2:0.1, 8:2:0.2, 7:3:0.5, 6:4:1, 0:1:0)] to
2
effect was measured by using the phenazine methosulfate
(PMS)–NADH system according to previously described
methods [14, 15]. Briefly, the reaction was started by the
addition of NADH into the assay mixture containing nitro
blue tetrazolium (NBT) plus varying concentrations of test
samples, then allowed to proceed for 10 min at room
temperature. The absorbance of the resulting solution was
measured at 570 nm. In both cases, Trolox was used as a
standard sample. Data shown represent mean ± SD
derived from four determinations.
afford 4 (9 mg), fr. 2.10.1 (20 mg), fr. 2.10.2 (7 mg), fr.
2
.10.3 (6 mg), and fr. 2.10.4 (13 mg).
1
1
1
: Amorphous powder. [a]D -64.1° (c 0.7, MeOH).
?
Positive FAB–MS m/z: 631 [M ? Na] . HR-positive
FAB–MS m/z: 631.1855 (calcd for C H O Na:
2
5
36 17
13
1
6
31.1850). H-NMR spectral data: see Table 1. C-NMR
spectral data: see Table 2.
: Amorphous powder. [a]D -23.3° (c 0.9, MeOH).
1
1
2
?
Positive FAB–MS m/z: 601 [M ? Na] . HR-positive
FAB–MS m/z: 601.2133 (calcd for C H O Na:
5
2
38 15
13
1
Acknowledgments We express our appreciation to Mr. H. Haraz-
ono of Fukuoka University for measurement of the FAB–MS. The
authors thank Mr. Shuhei Hasebe and Mr. Shinya Inaba for technical
assistance. This research was supported in part by a Grant-in-Aid for
Scientific Research (C) (No. 19590030) from Japan Society for the
Promotion of Science.
6
01.2108). H-NMR spectral data: see Table 1. C-NMR
spectral data: see Table 2.
: Amorphous powder. [a]D -22.6° (c 0.3, MeOH).
1
1
3
?
Positive FAB–MS m/z: 587 [M ? Na] . HR-positive
FAB–MS m/z: 587.1962 (calcd for C H O Na:
4
2
36 15
13
87.1952). H-NMR spectral data: see Table 1. C-NMR
1
5
spectral data: see Table 2.
References
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3
4
(
Showa Denko, Tokyo, 150 mm 9 6.0 mm); solvent,
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3
2
perature, 70°C; detector, JASCO OR-2090 plus (JASCO);
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R
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L-arabinose [tR (min) 5.8; optical activity, positive] and
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7
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