Ovafolinins A-E, five new lignans from Lyonia ovalifolia

Article abstract of DOI:10.1248/cpb.58.191

Five new lignans, ovafolinins A-E (1-5), were isolated from the wood of Lyonia ovalifolia (Ericaceae). The structures of 1-5 were elucidated based on 2D NMR spectroscopy, X-ray crystallography, and other chemical methods.

Full text of DOI:10.1248/cpb.58.191

February 2010
Chem. Pharm. Bull. 58(2) 191—194 (2010)
191
Ovafolinins A—E, Five New Lignans from Lyonia ovalifolia
a
a
a
Kenji KASHIMA, Kaichi SANO, Young Sook YUN, ,a Hiroji INA,b Akira KUNUGI,a and Hideshi INOUE
*
a School of Life Sciences, Tokyo University of Pharmacy and Life Sciences; and b School of Pharmacy, Tokyo University of
Pharmacy and Life Sciences; 1432–1 Horinouchi, Hachioji, Tokyo 192–0392, Japan.
Received September 2, 2009; accepted November 10, 2009; published online November 10, 2009
Five new lignans, ovafolinins A—E (1—5), were isolated from the wood of Lyonia ovalifolia (Ericaceae). The
structures of 1—5 were elucidated based on 2D NMR spectroscopy, X-ray crystallography, and other chemical
methods.
Key words Lyonia ovalifolia; Ericaceae; lignan; benzoxepin structure
Lignans are a widely distributed class of dimeric phenyl- groups (d 3.23, 3.77, 3.86, 4.06) and two aromatic protons (d
propanoids, and are one of the major classes of phytoestro- 6.26, 6.52) (Table 1). The 13C-NMR spectrum showed signals
gens1) that are known to alleviate menopausal symptoms and for four methoxyl carbons (d 55.8, 56.0, 59.1, 60.6), two
lower the risk of cardiovascular disease.2) Furthermore, they methylene carbons (d 69.3, 72.5), six methine (d 37.4, 39.7,
also possess anticancer3) and antiviral properties,4) inhibit 43.0, 79.0, 100.7, 104.8), and ten quaternary carbons
certain enzymes5) and exhibit antioxidant activity.6) In the (d 122.9, 124.5, 128.7, 135.0, 138.6, 143.9, 144.9, 145.7,
1
present study, we targeted lignans from Lyonia ovalifolia var. 146.1, 152.0) (Table 2). The H–1H correlation spectroscopy
elliptica, a deciduous tree distributed in Taiwan, China and (COSY), heteronuclear multiple quantum correlation
Japan. Yasue et al. identified lyoniols-A, -B, and -C to be (HMQC), and heteronuclear multiple bond correlation
toxic components isolated from the leaves,7) and lyoniside, (HMBC) spectra suggested 1 was an aryltetralin-type lignan.
which is a major component and lyoniresinol in the bark of The 1H–1H correlations were observed between H-7 and H-8,
this plant.8,9) In addition, Sakakibara et al. identified triter- H-8 and H-9, H-8 and H-8ꢂ, H-7ꢂ and H-8ꢂ, and H-8ꢂ and H-
pene glycosides in this plant.10,11) Herein, we report on five 9ꢂ. The strong HMBC correlations of H-1 with C-2, C-3, C-
novel lignans, ovafolinins A (1)—E (5), isolated from the 5, C-6 and C-7 and of H-7ꢂ with C-4 suggests that 1 contains
wood of L. ovalifolia. Furthermore, we have identified the A ring of an aryltetralin-type lignan. The existence of a B
ovafolinins A (1)—C (3) to bear a unique benzoxepin struc- ring was confirmed by the HMBC correlations of H-7 with
ture.
C-1, C-5, C-6, C-8, C-9 and C-9ꢂ and of H-7ꢂ with C-1ꢂ, C-
8ꢂ, and C-9ꢂ. The C ring was suggested by the HMBC corre-
lations of H-3ꢂ with C-1ꢂ, C-2ꢂ, and C-5ꢂ (Fig. 2). One of
Results and Discussion
The filtrate of the aqueous EtOH extract of L. ovalifolia af- methoxyl groups is linked to C-4ꢂ based on the HMBC cor-
forded new five lignans named ovafolinin A (1), B (2), C (3), relations of the methoxyl protons (d 3.23) with C-4ꢂ (d
D (4), and E (5) along with lyoniside (6) and lyoniresinol (7) 144.9). Another methoxyl group is connected to C-2 based
as known compounds. The ovafolinins were isolated by suc- on the HMBC correlation of H-1 and the methoxyl protons
cessive column chromatography on highly porous synthetic (d 3.86) with C-2 (d 146.1). The HMBC correlations of the
resin (Diaion HP-20), subsequent silica gel chromatography, third methoxyl protons (d 3.77) indicated connection to C-6ꢂ
and finally preparative reversed-phase HPLC.
(d 145.7). The position of the fourth methoxyl group is sug-
Ovafolinin A (1) was obtained as a colorless prisms. The gested to be at C-4 by HMBC correlations of methoxyl pro-
molecular formula was determined to be C22H24O8 by HR- tons (d 4.06) and H-7ꢂ with C-4 (d 143.9). These methoxyl
electrospray ionization (ESI)-MS, which showed a [MꢀH]ꢀ group assignments were confirmed by NOE correlations of
peak at m/z 417.1523 (Calcd for C22H25O8, 417.1549). The H-1 with the methyl protons of OCH3 at the positions 2, 4ꢂ,
IR spectrum showed absorptions for hydroxyl groups and 4, respectively. HMBC correlations of H-9 with C-2ꢂ and
(3392 cmꢁ1) and aromatic rings (1615 cmꢁ1). The UV ab- of H-9ꢂ with C-7, and the 13C-NMR chemical shifts indicate
sorption maxima occurred at 243 and 283 nm, implying the the existence of a methyleneoxy bridge between C-7 and C-
presence of conjugated double bond systems in the molecule. 9ꢂ and an ether bonding between C-2ꢂ and C-9. Based on the
1
The H-NMR spectrum showed signals for four methoxyl weak NOE correlations detected between H-1 and H-7, H-7
Fig. 1. Structures of Ovafolinins A (1)—E (5)
© 2010 Pharmaceutical Society of Japan
∗ To whom correspondence should be addressed. e-mail: yun@toyaku.ac.jp

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Vol. 58, No. 2
Table 1. 1H-NMR (500 MHz) Spectral Data for 1—5 at 300 Ka)
1b)
2b)
3c)
7.34 (1H, s)
4b)
5c)
1
7
6.52 (1H, s)
4.75 (1H, br d, 4.4)
6.38 (1H, s)
6.63 (1H, s)
7.51 (1H, s)
6.99 (1H, s)
2.86 (1H, br d, 17.4)
3.03 (1H, dd, 7.1, 17.4)
2.24 (1H, *)
4.82 (1H, d, 5.4)
8
9
2.32 (1H, *)
2.76 (1H, *)
3.10 (1H, *)
3.96 (1H, dd, 2.8, 13.3)
4.51 (1H, *br d, 13.3)
3.82 (1H, *)
4.41 (1H, dd, 2.7, 12.1)
3.90 (1H, *)
4.54 (1H, dd, 3.3, 11.9)
3.48 (1H, br d, 9.3)
4.24 (1H, dd, 6.4, 9.3)
7.10 (1H, d, 1.4)
9.49 (1H, s)
6.31 (1H, s)
2ꢂ
3ꢂ
6ꢂ
7ꢂ
8ꢂ
9ꢂ
6.26 (1H, s)
6.25 (1H, s)
6.33 (1H, s)
6.43 (1H, d, 1.4)
6.31 (1H, s)
4.80 (1H, s)
3.21 (1H, dd, 4.5, 10.2)
3.12 (1H, t, 10.2)
3.48 (1H, dd, 4.5, 10.2)
3.72 (3H, s)
4.49 (1H, d, 2.3)
2.60 (1H, *)
3.73 (1H, br d, 8.5)
4.11 (1H, dd, 2.7, 8.5)
3.86 (3H, s)
4.59 (1H, br s)
2.22 (1H, *)
3.62 (1H, dd, 7.1, 10.6)
3.72 (1H, *)
3.81 (3H, s)
4.06 (3H, s)
4.98 (1H, br s)
2.37 (1H, *)
3.51 (2H, *)
2.70 (1H, br t, 6.2)
3.86 (1H, br d, 9.8)
3.99 (1H, dd, 6.2, 9.8)
3.93 (3H, s)
3.95 (3H, s)
3.33 (3H, s)
OMe-2
OMe-4
OMe-3ꢂ
OMe-4ꢂ
OMe-5ꢂ
OMe-6ꢂ
3.81 (3H, s)
3.93 (3H, s)
4.06 (3H, s)
3.96 (3H, s)
3.33 (3H, s)
3.23 (3H, s)
3.77 (3H, s)
3.42 (3H, s)
3.74 (3H, s)
3.47 (3H, s)
3.75 (3H, s)
3.76 (3H, s)
3.72 (3H, s)
a) J-Values are given in Hz in parentheses. b) In CDCl3. c) In CD3OD. * Multiplicity was not determined due to overlapping and/or broadening of the signals.
Table 2. 13C-NMR (125 MHz) Spectral Data for 1—5 at 300 K
1a)
2a)
3b)
4a)
5b)
1
2
3
4
5
6
7
8
9
1ꢂ
2ꢂ
3ꢂ
4ꢂ
5ꢂ
6ꢂ
7ꢂ
104.8
146.1
138.6
143.9
122.9
128.7
79.0
105.4
146.1
136.3
144.6
122.4
126.1
29.3
104.2
149.0
147.0
146.9
125.7
132.9
200.4
51.6
106.6
145.4
139.9
145.4
121.1
126.7
81.2
109.7
149.5
145.3
147.9
127.0
123.9
149.0
136.4
194.3
136.3
106.1
149.2
135.0
149.2
106.1
38.5
43.0
69.3
33.9
80.2
48.9
64.1
79.1
124.5
152.0
100.7
144.9
135.0
145.7
37.4
123.8
152.8
145.3
136.3
145.4
101.1
29.9
124.2
153.6
146.9
137.3
148.5
102.4
31.5
134.3
101.7
146.7
133.3
146.5
100.2
86.0
8ꢂ
39.7
43.4
46.8
58.4
43.6
9ꢂ
72.5
64.8
65.0
64.0
62.7
OMe-2
OMe-4
OMe-3ꢂ
OMe-4ꢂ
OMe-5ꢂ
OMe-6ꢂ
56.0
60.6
55.7
61.5
56.5
61.7
56.3
61.0
61.0
56.6
56.8
60.7
Fig. 3. X-Ray Crystallographic Structure of Ovafolinin A (1)
termined as shown in Fig. 1. This assignment was confirmed
by single-crystal X-ray diffraction analysis with a suitable
single crystal, obtained by careful crystallization from
methanol/water (Fig. 3).
59.1
55.8
59.6
55.9
60.2
56.5
56.3
56.6
a) In CDCl3. b) In CD3OD.
Ovafolinin B (2) was obtained as a colorless prism. Its mo-
lecular formula was determined to be C22H26O8 by HR-ESI-
MS [MꢀH]ꢀ m/z 419.1673 (Calcd for C22H27O8, 419.1706).
The IR and UV spectra of 2 were similar to those of 1. The
1H- and 13C-NMR spectra of 2 were also similar to those of
1, indicating the presence of four methoxyl groups (dH 3.42,
dC 59.6; dH 3.74, dC 55.9; dH 3.81, dC 55.7; dH 4.06, dC
61.5), two aromatic protons (dH 6.25, dC 101.1; dH 6.38, dC
105.4), and one oxymethylene carbon (dH 3.82 and 4.41, dC
1
80.2) (Tables 1, 2). The similarity in H–1H COSY and
Fig. 2. Key HMBC and NOE Correlations for Determination of the
Ovafolinin A (1)
HMBC spectra between 2 and 1 suggest that 2 has the same
basic structure as 1. Although the HMBC correlations be-
and H-8, H-8 and H-8ꢂ, and H-7ꢂ and H-8ꢂ (Fig. 2), the rela- tween H-7 and C-1, C-8 and C-9 and between H-7ꢂ and C-4,
tive stereochemistry was assigned to be 7R*, 8S*, 7ꢂR*, and C-5, C-6, C-1ꢂ, and C-2ꢂ showed that 2 is aryltetralin type
8ꢂR*. Accordingly, the structure of ovafolinin A (1) was de- lignan, similar to 1, a major difference between 2 and 1 was

February 2010
193
1
observed based on the H- and 13C-NMR chemical shifts at of hydroxyl groups (3270 cmꢁ1). Conjugated systems were
position 7. The proton chemical shifts were d 4.75 for 1 and also suggested by the UV absorption maximum at 276 nm
1
d 2.86 and 3.03 for 2 and the carbon chemical shifts were d (log e 3.07). The H- and 13C-NMR spectra of 4 were very
79.0 for 1 and d 29.3 for 2. This suggests that the oxymethin similar to those of 1, indicating the aryltetralin-lignan struc-
1
of 1 is replaced by a methylene in 2. The NOE correlations ture. The H- and 13C-NMR chemical shifts at position 7 (d
between H-1 and Ha-7, Ha-7 and H-8, and H-7ꢂ and H-8ꢂ of 2 4.82, 81.2) at position 9ꢂ (d 3.86/3.99, 64.1) and the HMBC
were similar to those of 1 (Fig. 4). Thus, the structure of 2 correlations of H-7 with C-9ꢂ and of H-9ꢂ with C-7, indi-
was determined as shown in Fig. 1.
cated an ether linkage was assigned between C-7 and C-9ꢂ,
Ovafolinin C (3) was obtained as a yellowish-white amor- similar to ovafolinin A (1). Furthermore, the HMBC correla-
phous solid. Its molecular formula was determined to be tions of H-9, H-2ꢂ, H-6ꢂ and H-8 with the quaternary carbon
C22H24O9 by HR-ESI-MS [MꢀH]ꢀ m/z 433.1465 (Calcd for C-7ꢂ (d 86.0) and the chemical shift of C-9 at d 64.1 and of
C22H25O9, 433.1499). The IR spectrum of 3 indicated the C-7ꢂ at d 86.0 demonstrated the presence of an ether linkage
presence of hydroxyl (3272 cmꢁ1) and ketone (1670 cmꢁ1) between C-9 and C-7ꢂ. The absolute configuration of
groups. The UV absorption maxima occurred at 243 and ovafolinin D (4) was established by comparison of ovafolinin
290 nm, suggesting the presence of conjugated double bonds D dimethyl ether (9) from ovafolinin D (4) and that prepared
1
in the molecule. The H- and 13C-NMR spectra of 3 were from lyoniside (6) as shown in Chart 1. Because the two de-
similar to those of 2, with the exception that there were only rivatives were indistinguishable in the NMR and MS data,
signals for position 7. The signals at d 2.86 and 3.03 in 2 the absolute structure of 4 was determined as shown in Fig.
were not detected in of the spectra 3. Furthermore the chemi- 1.
cal shift for C-7 of 3 was d 200.4 (in CD3OD), whereas that
Ovafolinin E (5) was obtained as a yellowish-white amor-
of 2 was d 29.3 (in CDCl3). The HMBC correlations ob- phous solid. Its molecular formula was determined to be
served of H-1, H-8ꢂ and H-8 with C-7, and the IR spectrum C22H24O8 by HR-ESI-MS [MꢀH]ꢀ m/z 417.1562 (Calcd for
indicate the presence of a ketone group. Thus, ovafolinin C C22H25O8 417.1549). The IR spectrum showed absorptions
(3) was assigned as shown in Fig. 1.
for hydroxyl (3306 cmꢁ1) and carbonyl groups (1660 cmꢁ1).
Ovafolinin D (4) was obtained as a yellowish-white amor- The UV absorption maxima occurred at 253 (log e 4.10) and
phous solid. Its molecular formula was determined to be 354 nm (log e 4.01), implying the presence of conjugated
C22H24O8 by HR-ESI-MS [MꢀH]ꢀ m/z 417.1582 (Calcd for double bond systems in the molecule. The 1H- and 13C-NMR
C22H25O8, 417.1549). The IR spectrum showed the presence spectra showed the signals attributed to an aldehyde group (d
9.49, 194.3) and olefinic carbons (d 149.0, 136.4). The alde-
hyde group was located at position 9 based on correlations
between H-7 and C-9, H-9 and C-8 (d 136.3) in HMBC
1
spectrum data. Analysis of the H–1H COSY, HMQC, and
HMBC spectra, ovafolinin E (5), unlike the others (1—4),
has no ether linkage between any of carbons in the arylte-
tralin-lignan skeleton (Fig. 1).
Ovafolinins A (1)—E (5) were found to be inactive in the
cytotoxic activity test on HL 60, HCT116, A549, and MCF7
cell lines.
Fig. 4. Selected NOE Correlations of Ovafolinin B (2)
Ovafolinins A (1)—C (3) were determined to be unique
Reagent; (a) CH3I/K2CO3, aceton, 5%H2SO4, (b) CuSO4/K2S2O8, CH3CN, (c) CH3I/K2CO4.
Chart 1. Derivation of Ovafolinin D Dimethyl Ether (8) from Lyoniside (6) and Ovafolinin D (4)

194
Vol. 58, No. 2
Oxidation of Lyoniresinol Dimethyl Ether (8)12,13) Lyoniresinol
dimethyl ether (8, 40 mg, 0.089 mmol, 14.4 ml CH3CN), CuSO4·5H2O
(24 mg, 0.15 mmol, 1.6 ml H2O) and K2S2O8 (51.6 mg, 0.19 mmol, 4.4 ml
H2O) were refluxed for 0.5 h at 120 °C, diluted with H2O, and extracted with
AcOEt. Ovafolinin D dimethyl ether (9, 2.9 mg) was isolated by HPLC
(CHCl3 : AcOEtꢄ3 : 1). Ovalifolin D dimethyl ether (9); Amorphous solid,
aryltetralin-lignans containing 7-membered ring formed by
the C9–C-2ꢂ ether linkage. 1—3 bear a benzoxepin structures
including the C ring of the aryltetralin-lingnan. In addition,
ovafolinin D (4) is the first example of a naturally occurring
aryltetralin-lignan bearing methyleneoxy-bridge.
[a]D25 ꢁ60° (cꢄ0.05, MeOH). H-NMR (CDCl3) d: 7.07 (1H, d, Jꢄ1.7 Hz,
1
Experimental
H-2ꢂ), 6.61 (1H, s, H-6), 6.39 (1H, d, Jꢄ1.7 Hz, H-6ꢂ), 4.83 (1H, d,
Jꢄ5.4 Hz, H-7), 4.25 (1H, dd, Jꢄ6.4, 9.5 Hz, H-9a), 4.04 (1H, dd, Jꢄ6.3,
9.8 Hz, H-9ꢂa), 3.93 (1H, overlapped, H-9ꢂb), 3.92 (3H, s, H-3ꢂ), 3.90 (3H, s,
H-3), 3.86 (3H, s, H-4ꢂ), 3.76 (3H, s, H-4), 3.73 (3H, s, H-3ꢂ), 3.46 (1H, dd,
Jꢄ8.5, 9.5 Hz, H-9b), 3.32 (3H, s, H-5ꢂ), 3.13 (1H, dd, Jꢄ6.4, 9.5 Hz, H-8),
2.73 (1H, dd, Jꢄ6.3, 6.1 Hz, H-8ꢂ). 13C-NMR (CDCl3) d: 154.0 (C-3), 153.2
(C-3ꢂ), 152.5 (C-5ꢂ), 152.4 (C-5), 143.4 (C-4), 139.7 (C-1ꢂ), 136.4 (C-4ꢂ),
131.0 (C-1), 121.1 (C-2), 106.7 (C-6), 101.2 (C-6ꢂ), 100.2 (C-2ꢂ), 85.9 (C-
7ꢂ), 81.0 (C-7), 64.3 (C-9), 64.0 (C-9ꢂ), 60.8 (–OCH3), 60.6 (–OCH3ꢃ2),
57.9 (C-8ꢂ), 56.1 (–OCH3), 56.0 (–OCH3), 55.9 (–OCH3), 49.0 (C-8). HR-
ESI-MS m/z: 445.1860 [MꢀH]ꢀ (Calcd for C24H29O8Na, 445.1862).
Methylation of Ovafolinin D (4) CH3I (1 ml) and anhydrous K2CO3
(8 mg) were added to a solution 4 (1.2 mg, 0.0029 mmol) in acetone
(0.04 ml) while stirring. Then the mixture maintained with stirring for 30 h
at room temperature. After evaporation and purification of the crude product
by HPLC (CHCl3 : AcOEtꢄ3 : 1) afforded the methylated product (0.5 mg).
1H-NMR (CDCl3) d: 7.08 (1H, d, Jꢄ1.7 Hz, H-2ꢂ), 6.62 (1H, s, H-6), 6.39
(1H, d, Jꢄ1.7 Hz, H-6ꢂ), 4.82 (1H, d, Jꢄ5.3 Hz, H-7), 4.26 (1H, dd, Jꢄ6.3,
9.7 Hz, H-9a), 4.04 (1H, dd, Jꢄ6.3, 9.7 Hz, H-9ꢂa), 3.93 (1H, overlapped, H-
9ꢂb), 3.93 (3H, s, H-3ꢂ), 3.90 (3H, s, H-3), 3.86 (3H, s, H-4ꢂ), 3.74 (3H, s,
H-4), 3.73 (3H, s, H-3ꢂ), 3.46 (1H, overlapped, H-9b), 3.33 (3H, s, H-5ꢂ),
3.12 (1H, dd, Jꢄ6.3, 9.5 Hz, H-8), 2.73 (1H, dd, Jꢄ6.3, 6.1 Hz, H-8ꢂ). HR-
ESI-MS m/z: 445.1882 [MꢀH]ꢀ (Calcd for C24H29O8Na, 445.1862).
General Procedures Optical rotations were measured on a JASCO
DIP-1000 digital polarimeter. IR spectra were recorded on a FT-IR 1710
spectrophotometer. UV spectra were obtained using a Hitachi U-2001 spec-
trophotometer. NMR spectra were measured on Bruker DRX-500 and DPX-
400 spectrometers at 300 K. The 1H-NMR chemical shifts in CDCl3 and
CD3OD calibrated to the residual CHCl3 and CH3OH resonances at 7.26 and
3.31 ppm, respectively, and the 13C-NMR chemical shifts were calibrated to
the solvent peaks at 77.0 and 49.0 ppm, respectively. Mass spectra were ob-
tained using a Micromass LCT spectrometer. Preparative HPLC was carried
out on a Shimadzu LC-6AT system equipped with a SPD-10AVP detector
and a reversed-phased column, Mightysil RP-18 prep (5 mg, 20ꢃ250 mm),
using CH3OH : H2O or a CH3CN : H2O as the mobile phase, at a flow rate of
5 ml/min.
Plant Collection Wood from L. ovalifolia was collected in Saitama Pre-
fecture, in October 2000, and the plant origin was identified by Dr. H. Ina
(Tokyo University of Pharmacy and Life Sciences, Japan).
Extraction and Isolation Fresh wood chips (15 kg) from L. ovalifolia
were extracted using MeOH. After removal of MeOH in vacuo, the residue
was dissolved in water. The aqueous layer was separated from the precipi-
tates, and placed on a HP-20 column (DIAION). Elution with H2O/MeOH
mixtures (100 : 0, 80 : 20, 60 : 40, 40 : 60, 20 : 80 and 0 : 100) and acetone af-
forded seven fractions (frs. Lo 1—7). Fraction Lo 5 (26.1 g) was subjected
to silica gel (Merck Kieselgel 60) column chromatography eluting sequen-
tially with CHCl3/MeOH mixture (50 : 1, 20 : 1, 10 : 1, 7 : 3, and 1 : 1). The
CHCl3/CH3OH (50 : 1) fraction (681.7 mg) was evaporated and applied to
ODS HPLC eluting with CH3CN : H2O (60 : 40) to afford 1 (37.8 mg) and 5
(3.6 mg), and with CH3CN : H2O (70 : 39) to afford 2 (5.2 mg), 3 (3.2 mg)
and 4 (3.7 mg).
X-Ray Crystallographic Study of Ovalifolin
A (1) C22H24O8,
Mꢄ416.41, 0.49ꢃ0.45ꢃ0.38. Single-crystal X-ray analysis was carried out
on a Mac Science DIP diffractometer with MoKa radiation (lꢄ0.71073).
The data indicated the monoclinic space group, Cc, aꢄ19.6130 (19) Å,
bꢄ7.0010 (7) Å, cꢄ15.2280 (8) Å, Vꢄ1955.5 (3) Å3, Zꢄ4, Dxꢄ1.414
Mg mꢁ3, 2081 measured reflections, 2081 independent reflections, 1589 ob-
served reflections [Iꢅ2s(I)], R1ꢄ0.0387, wR2ꢄ0.0899 (observed data),
GOFꢄ0.930; R1ꢄ0.0476, wR2ꢄ0.0925 (all data). The structure was solved
by direct methods using the maXus crystallographic software package, and
refined by full-matrix least-squares on F2 using the program SHELXL-97.
Crystallographic data for the structure of 1 have been deposited in the Cam-
bridge Crystallographic Data Centre under deposition number CCDC
722340.
Ovafolinin A (1): Colorless prisms, [a]D25 ꢁ37.3° (cꢄ0.36, MeOH). IR
(neat) cmꢁ1: 3392, 2940, 1615. UV lmax (MeOH) nm (log e): 243 (3.86),
283 (3.58). 1H- and 13C-NMR spectrometric data are given in Tables 1 and 2,
respectively. HR-ESI-MS m/z: 417.1523 [MꢀH]ꢀ (Calcd for C22H25O8,
417.1549).
Ovafolinin B (2): Colorless prisms, [a]D25 ꢀ52.0° (cꢄ0.26, MeOH). IR
(neat) cmꢁ1: 3370, 2936, 1613. UV lmax (MeOH) nm (log e): 283 (3.99) nm.
1H- and 13C-NMR spectrometric data are given in Tables 1 and 2, respec-
tively. HR-ESI-MS m/z: 419.1673 [MꢀH]ꢀ (Calcd for C22H27O8 419.1706).
Ovafolinin C (3): Amorphous solid, [a]D25 ꢀ105.7° (cꢄ0.11, MeOH). IR
(neat) cmꢁ1: 3272, 2938, 1567. UV lmax (MeOH) nm (log e): 290 (3.85).
1H- and 13C-NMR spectrometric data are given in Tables 1 and 2, respec-
tively. HR-ESI-MS m/z: 433.1465 [MꢀH]ꢀ (Calcd for C22H25O9 433.1499).
Ovafolinin D (4): Amorphous solid, [a]D25 ꢁ33° (cꢄ0.09, MeOH). IR
(neat) cmꢁ1: 3389, 2930, 1614. UV lmax (MeOH) nm (log e): 276 (3.07).
1H- and 13C-NMR spectrometric data are given in Tables 1 and 2, respec-
tively. HR-ESI-MS m/z: 417.1582 ([MꢀH]ꢀ (Calcd for C22H25O8 417.1549).
Ovafolinin E (5): Amorphous solid. [a]D25 ꢀ105.7° (cꢄ0.16, MeOH); IR
(neat) cmꢁ1: 3272, 2938, 1567. UV lmax (MeOH) nm (log e): 290 (3.85).
1H- and 13C-NMR spectrometric data are given in Tables 1 and 2, respec-
tively. HR-ESI-MS m/z: 417.1562 [MꢀH]ꢀ (Calcd for C22H25O8 417.1549).
Conversion of Lyoniside (6) to Lyoniresinol Dimethyl Ether (8) Ly-
oniside (6, 1 g, 1.8 mmol) and CH3I (1 ml) were stirred in acetone (40 ml) at
room temperature for 66 h to afford lyoniside dimethyl ether (330 mg) by
silica gel chromatography (CHCl3 : MeOHꢄ10 : 1). This product was dis-
solved in EtOH (10 ml) and treated with an aqueous solution 5% H2SO4
(2 ml) at 110 °C under reflux for 7 h. After cooling, H2O was added to the
mixture, and it was extracted with AcOEt. The AcOEt layer was dried over
Mg2SO4, evaporated, and subjected to silica gel column chromatography
(CHCl3 : MeOHꢄ20 : 1). The product was isolated and crystallized from
benzene to yield lyoniresinol dimethyl ether (8, 208 mg). Lyoniresinol di-
methyl ether: HR-ESI-MS [MꢀNa]ꢀ m/z: 471.2015 (Calcd for C24H32O8Na,
471.2019).
Acknowledgements The authors are grateful to Prof. H. Morita and Dr.
T. Hosoya for cytotoxicity assay.
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