Oxylipins arabidopsides C and D from Arabidopsis thaliana

Article abstract of DOI:10.1021/np0495938

Two new oxylipins, arabidopsides C (1) and D (2), were isolated from the aerial parts of Arabidopsis thaliana, and the structures of 1 and 2 were elucidated using spectroscopic data, primarily NMR and MS, and chemical means. Arabidopsides C (1) and D (2) are rare digalactosyl diacylglycerides containing 12-oxophytodienoic acid and/or dinor-oxophytodienoic acid. Arabidopside D (2) and arabidopsides A (3) and B (4), which were also isolated from this plant, exhibited inhibitory effects on the growth of the root of cress (Lepidium sativum) seedlings at 5 × 10^-5 mol/L.

Full text of DOI:10.1021/np0495938

6
00
J. Nat. Prod. 2005, 68, 600-603
Oxylipins Arabidopsides C and D from Arabidopsis thaliana
†
‡
§
†
,†
Yosuke Hisamatsu, Nobuharu Goto, Mitsuhiro Sekiguchi, Koji Hasegawa, and Hideyuki Shigemori*
Graduate School of Life and Environmental Sciences, University of Tsukuba, Tsukuba 305-8572, Japan,
Department of Biology, Miyagi University of Education, Sendai 980-0845, Japan, and Institute for Drug Discovery Research,
Yamanouchi Pharmaceutical Co., Ltd., Tsukuba 305-8585, Japan
Received December 17, 2004
Two new oxylipins, arabidopsides C (1) and D (2), were isolated from the aerial parts of Arabidopsis
thaliana, and the structures of 1 and 2 were elucidated using spectroscopic data, primarily NMR and
MS, and chemical means. Arabidopsides C (1) and D (2) are rare digalactosyl diacylglycerides containing
1
2-oxophytodienoic acid and/or dinor-oxophytodienoic acid. Arabidopside D (2) and arabidopsides A (3)
and B (4), which were also isolated from this plant, exhibited inhibitory effects on the growth of the root
-
5
of cress (Lepidium sativum) seedlings at 5 × 10 mol/L.
Arabidopsis thaliana ecotype col-0 (Brassicaceae) is
known as a model plant, and biologists have used this plant
for various genetic studies. However, few examples of
bioactive substances from A. thaliana have been reported,
except for some phytohormones. We previously isolated two
unique oxylipins, arabidopsides A (3) and B (4), from the
1
aerial parts of A. thaliana. Compounds 3 and 4 are rare
monogalactosyl diacylglycerides containing 12-oxophyto-
dienoic acid (OPDA) and/or dinor-oxophytodienoic acid (dn-
OPDA), which are known as precursors of jasmonic acid
2
,3
(
JA).
JA and OPDA have received much attention
because they play important roles in regulation of devel-
4
opmental and defense gene expression of plants. Further
examination of an extract of A. thaliana resulted in
isolation of two additional oxylipins, arabidopsides C (1)
and D (2), which are new digalactosyl diacylglycerides
containing OPDA and/or dn-OPDA. This paper describes
the isolation and structure elucidation of 1 and 2 and their
inhibitory effect on cress root growth.
The aerial parts (100 g) of A. thaliana were extracted
with MeOH. The MeOH extract was partitioned between
EtOAc and H
to a silica gel column (CHCl
glycolipid fraction, which was fractionated by reversed-
phase C18 HPLC (CH CN/H O, 1:1) to give arabidopsides
2
O. The EtOAc-soluble portion was subjected
3
/MeOH, 1:1) to afford a
3
2
C (1, 0.0004%, wet weight) and D (2, 0.0018%) as colorless
amorphous solids, together with arabidopsides A (3,
0
.0040%) and B (4, 0.0018%).1
The molecular formula, C51
H
80
O
17, of arabidopside D (2)
+
was established by HRESIMS [m/z 987.5285 (M + Na) ,
∆
-0.8 mmu]. The IR spectrum implied the presence of
-
1
-1
hydroxy (3424 cm ), ester carbonyl (1736 cm ), and
-
1
unsaturated carbonyl (1714 and 1631 cm ) groups. The
gross structure of 2 was deduced from detailed analysis of
1
13
the H and C NMR data (Table 1) aided by 2D NMR
1
1
1
experiments ( H- H COSY, HMQC, and HMBC). The H
1
3
and C NMR data of 2 were similar to those of arabidop-
side B (4) except for signals due to the sugar moieties. The
1
1
H- H COSY connectivities of C-1 through C-3, C-1′
H-1′ to H-3′, H-4′ to H-3′ and H-5′, H-1′′ to H-3′′, and H-4′′
to H-3′′ and H-5′′ and the H coupling constants (Table
through C-6′, and C-1′′ through C-6′′ indicated the presence
of a glycerol and two sugar components. The two sugars
were assigned to be galactoses by NOESY correlations of
1
H-
1
1). The anomeric proton (H-1′, δ 4.29) of 2 showed HMBC
H
cross-peaks with C-3 (δ 68.6) and C-2′ (δ 73.1), and the
C
C
other anomeric proton (H-1′′, δ
H
C
4.77) with C-6′ (δ 68.6)
1
13
and C-2′′ (δ
C
71.0). The H and C NMR spectroscopic data
*
To whom correspondence should be addressed. Tel & Fax: +81-29-
8
53-4603. E-mail: hshige@agbi.tsukuba.ac.jp.
and H- C coupling constants obtained by a nondecoupled
HSQC experiment of 2 in CD OD (C-1′, JCH ) 169.8 Hz;
3
1
13
†
University of Tsukuba.
‡
Miyagi University of Education.
§
Yamanouchi Pharmaceutical Co., Ltd.
C-1′′, JCH ) 162.8 Hz) revealed that 2 possessed the 1,2-
1
0.1021/np0495938 CCC: $30.25
© 2005 American Chemical Society and American Society of Pharmacognosy
Published on Web 03/26/2005

Notes
Journal of Natural Products, 2005, Vol. 68, No. 4 601
Table 1. 13C and H NMR Assignments (δ) for the Glycerol
1
and Sugar Moieties of 1 and 2
arabidopside C (1)
arabidopside D (2)
13Ca
1Hb
13Ca
1Hb
1
64.5 4.44 (dd, 5.4, 10.8)
64.9 4.26 (dd, 6.6, 12.0)
4.47 (dd, 2.4, 12.0)
4.24
m
m
m
m
2
3
71.1 5.29
67.9 4.00
71.4 5.29
m
68.6 3.98 (dd 5.4, 10.8)
3
.76
3.76
m
1′
2′
3′
4′
5′
6′
106.1 4.27 (d, 7.2)
106.0 4.29 (d, 7.2)
73.2 3.55 (dd, 7.2, 14.1)
75.6 3.51 (dd, 4.2, 14.1)
71.0 3.92 (d, 4.2)
73.1 3.53 (dd, 7.8, 12.6)
75.5 3.52 (dd, 2.4, 10.2)
71.0 3.92 (d, 2.4)
75.5 3.78
67.1 3.96
m
m
m
75.4 3.78
m
68.6 3.72 (dd, 4.8, 11.4)
3.71
3.93
m
1
2
3
4
5
6
′′ 101.4 4.91 (d, 3.2)
101.2 4.77 (d, 3.6)
′′
′′
′′
′′
′′
70.8 3.82 (dd, 3.2, 10.2)
72.2 3.76 (dd, 3.3, 10.2)
71.9 3.94 (d, 3.3)
71.0 3.83 (dd, 3.6, 10.2)
72.1 3.77 (dd, 3.0, 10.2)
71.8 3.94 (d, 3.0)
73.4 3.88
62.8 3.76
m
m
m
73.3 3.89
63.5 3.76
3.76
m
m
m
Figure 1. Derivatization of 1 and 2.
3.76
a
00 MHz; CD3OD (δ 49.8). b 600 MHz; CD3OD (δ 3.35).
6
di-O-acyl-3-O-[R-D-galactopyranosyl-(1′′f6′)-O-â-D-galacto-
5
1
1
syl]-sn-glycerol moiety. The H- H COSY connectivities
of C-9′′′ (or C-9′′′′) through C-11′′′ (or C-11′′′′) and C-9′′′ (or
C-9′′′′) through C-13′′′ (or C-13′′′′) and HMBC correlations
of H-11′′′ (or H-11′′′′) (δ
C-12′′′ (or C-12′′′′) (δ
9.8) indicated the presence of two cyclopentenone moieties.
C
6.21) to C-9′′′ (or C-9′′′′) (δ
C
44.8),
C
215.2), and C-13′′′ (or C-13′′′′) (δ
C
4
1
1
The H- H COSY connectivities of C-13′′′ (or C-13′′′′)
through C-18′′′ (or C-18′′′′) and HMBC correlations of Ha-
1
2
4′′′ (or Ha-14′′′′) (δ
.24) to C-15′′′ (or C-15′′′′) (δ
135.7) revealed that 2-penetene groups connected to
H
H
(
δ
C
Figure 2. Growth inhibitory effects of arabidopsides A (3), B (4), and
D (2).
C-13′′′ and C-13′′′′. Z-Geometries of two disubstituted
double bonds at C-15′′′-C-16′′′ and C-15′′′′-C-16′′′′ were
deduced from the carbon chemical shifts of allylic carbons
Therefore, arabidopside C (1) was assigned to be sn1-O-
(12-oxophytodienoyl)-sn2-O-(dinor-oxophytodienoyl)digalac-
tosyl diglyceride.
6
(
C-14′′′ or C-14′′′′, δ
C
24.7; C-17′′′ or C-17′′′′, δ
C
22.3). These
data and proton and carbon resonances indicated that 2
possessed two cis-12-oxophytodienoic acids (OPDA). HMBC
correlations of Ha-1 and Hb-1 to an ester carbonyl carbon
As JA showed inhibitory effects on root growth of various
4
plants, the effects of arabidopsides A (3), B (4), and D (2)
(
δ
C
176.8) and chemical shifts (δ
H
5.29; δ
C
71.4) of C-2
were also examined by the application to cress seeds. Ten
seeds of cress (Lepidium sativum L.) were placed on a filter
paper moistened with test solution and kept for 40 h at 24
°C in the dark, after which the lengths of their roots were
measured. As shown in Figure 2, arabidopsides A (3), B
(4), and D (2) inhibited 30% of the growth of cress roots at
indicated that the OPDA connected to C-1 and C-2. The
chiral GC analysis of methanolisates of 2 after treatment
with HCl/MeOH detected the mixture (10:3) of (9S,13S)-5
7
and (9R,13R)-5 (Figure 1). On the other hand, the dibenzyl
glycerol (6) derived from 2 was racemic by chiral GC
analysis (Figure 1).8
-5
5 × 10 mol/L, while OPDA and JA inhibited root growth
-
5
Arabidopside C (1) showed a pseudomolecular ion peak
50% at 5 × 10 mol/L.
+
at m/z 959 (M + Na) in the ESIMS. HRESIMS analysis
Arabidopsides C (1) and D (2) are the first digalactosyl
diacylglycerides containing OPDA and/or dn-OPDA, while
arabidopsides A (3) and B (4) and MGDG-O are mono-
galactosyl diacylglycerides containing OPDA and/or dn-
revealed the molecular formula of 1 to be C49
76
H O17 [m/z
+
9
59.4971 (M + Na) , ∆ -0.9 mmu], indicating that 1 was
1
13
an ethylene homologue of 2. The H and C NMR spectra
of 1 were similar to those of 2 except for the signals due to
the methylene chains. The two lipids were presumed to be
OPDA and dn-OPDA moieties judging from spectral data
of OPDA and dn-OPDA in arabidopside A (3).1 HMBC
1,11
OPDA.
Arabidopsides A (3), B (4), and D (2) exhibited
-5
inhibitory effects on the growth of cress roots at 5 × 10
mol/L, suggesting that the sugar moiety has little effect
on growth activity.
correlations of Ha-1 (δ
carbonyl carbon (δ 175.1) and chemical shifts (δ
1.1) of C-2 indicated that OPDA and dn-OPDA connected
H
4.44) and Hb-1 (δ
H
4.24) to the ester
C
H
5.29; δ
C
Experimental Section
7
General Experimental Procedures. Optical rotations
to C-1 and C-2. To define the locations of these lipids in
the â-galactosylglycerol moiety of 1, we applied enzymatic
hydrolysis with lipase type XI (Sigma) to preferentially
were measured with a JASCO DIP-370 polarimeter. IR spectra
1
were recorded on a JASCO FT/IR-300 spectrometer. H and
13
C NMR spectra were measured and recorded on a Varian
9
hydrolyze sn1 fatty acids. The lipase type XI-catalyzed
Unity INOVA 600 spectrometer in CD OD. The resonances of
3
hydrolysis of 1 afforded OPDA (7, Figure 1), which was
identified as OPDA methyl ester derived from the hydroly-
sate with trimethylsilyldiazomethane,10 using GC analysis.
CD OD at δ 3.35 ppm and δ 49.8 ppm were used as internal
references for NMR spectra. ESIMS were recorded on a
micromass Q-Tof-2 mass spectrometer.
3
H
C

6
02 Journal of Natural Products, 2005, Vol. 68, No. 4
Notes
Plant Material. The seeds of Arabidopsis thaliana ecotype
(OPDAMe, 5) were determined in comparison to the retention
time (min) of OPDAMe derived from OPDA, purchased from
Cayman Chemical, by methylation with trimethylsilyldiazo-
col-0 (Brassicaceae) were purchased from Lehle Seeds. The
2
seeds were immersed in H O at 4 °C for 2 days before sowing
1
0
on rock wool (rock fiber, NITTOBO, Japan). They were then
cultured under continuous light (24 h, ca. 3800 lux) at 24 °C,
until forming a flower bud.
methane. The cis enantiomers, (9′′′S, 13′′′S)-5 (tR 397 min)
and (9′′′R,13R′′′)-5 (tR 397 min), (9′′′′S,13S′′′′)-5′ (tR 397 min)
and (9′′′′R,13R′′′′)-5′ (t 397 min), were inseparable. The trans
R
Extraction and Isolation. Aerial parts of A. thaliana (100
g) were extracted with MeOH (1 L × 2) and evaporated to
dryness in vacuo at 30 °C. The MeOH extract (3.66 g) was then
enantiomers, (9′′′S,13′′′R)-5 (t 347 min) and (9′′′R,13′′′S)-5 (t
R
R
3
3
50 min), (9′′′′S,13′′′′R)-5′ (t
R
347 min) and (9′′′′R,13′′′′S)-5′ (t
R
50 min) (10:3), were produced from cis forms by enolization.
partitioned between EtOAc (100 mL × 3) and H
The EtOAc-soluble portion (0.9 g) was subjected to a silica gel
column (1.1 × 31 cm, CHCl /MeOH, 19:1 f 1:1). The fraction
eluted with CHCl /MeOH, 1:1, was applied to a C18 Sep-Pak
cartridge (CH CN/H O, 3:1, Waters), and the fraction (10.3 mg)
2
O (100 mL).
1
, 2-Dibenzyl Glycerol (6) Derived from 2 and Chiral
GC Analysis of 6. This derivatization followed the method
3
8
of Uzawa. A solution of 2 (1.2 mg) in dry MeOH (0.1 mL) was
3
treated with NaOMe/MeOH (10 equiv) at room temperature
for 1 h. The reaction mixture was partitioned between hexane
3
2
containing arabidopsides C (1) and D (2) was further separated
by reversed-phase HPLC [Deverosil ODS HG-5 (Nomura
Chemical, φ 1.0 × 25 cm), flow rate 2.5 mL/min; solvent
and H
2
2 2
O, and the H O-soluble portion was concentrated under
N
gas. A solution of the concentrate in dry DMF (1 mL) was
treated with NaH at room temperature for 30 min. Benzyl
bromide (5 µL) was added to the reaction solution, then the
mixture was stirred at room temperature for a further 14 h.
The reaction was quenched by the addition of saturated
3 2
CH CN/H O (11:9); detection UV (222 nm)] to give arabidop-
sides C (1, 1.8 mg, t
The CHCl /MeOH, 19:1, fraction was separated by reversed-
phase HPLC [Deverosil ODS HG-5, flow rate 2.5 mL/min;
R R
22 min) and D (2, 0.4 mg, t 38 min).
3
4
aqueous NH Cl and extracted with EtOAc (5 mL) three times.
3 2
solvent CH CN-H O (4:1); detection UV (222 nm)] to afford
arabidopsides A (3, 4.0 mg, t 10 min) and B (4, 1.8 mg, t 13
R R
The EtOAc extract was concentrated in vacuo. The residual
syrup was subjected to preliminary separation by TLC [benzene/
hexane (10:1) f EtOAc], and a solution of the concentrate in
min).
2
4
Arabidopside C (1): colorless amorphous solid; [R]
30.0° (c 0.69, MeOH); IR (KBr) νmax 3424, 1736, 1714, and
D
1
0% dry HCl/MeOH (600 µL) was heated at 100 °C for 8 h.
The reaction mixture was poured into EtOH and concentrated
under N gas. The reaction mixture (0.19 mg) was subjected
+
1
-
1 1
3
631 cm ; H NMR (600 MHz, CD OD) δ 7.96 (2H, m, H-8′′′,
H-10′′′′), 6.20 (2H, m, H-9′′′, H-11′′′′), 5.45 (4H, m, H-13′′′,
H-14′′′, H-15′′′′, H-16′′′′), 3.10 (2H, m, H-7′′′, H-9′′′′), 2.54 (2H,
m, H-11′′′, H-13′′′′), 2.48 (2H, m, Ha-12′′′, Ha-14′′′′), 2.39 (4H,
2
to chiral-GC analysis. Chiral-GC conditions: column: CP-
cyclodex-â-2,3,6 M-capillary column, Varian; program rate: 40
f 150 °C (at 10 °C/min), 150 °C (450 min), 150 f 200 °C (at
10 °C/min), and 200 °C (60 min). The absolute configuration
of the dibenzyl glycerol were determined by the retention time
(min) of (S)- and (R)-dibenzyl glycerol (S: 373 min, R: 376
min) derived from (S)- and (R)-glycerol acetonide according to
m, H
2
-2′′′, H
-15′′′, H
-3′′′, H -3′′′′), 1.36 (12H, m, H
′′′′, H -6′′′′, H -7′′′′), 1.27 (2H, m, Hb-8′′′, Hb-8′′′′), and 1.02
6H, t, J ) 7.2 Hz, H
CD OD) δ 213.8 (C-10′′′, C-12′′′′), 175.1 (C-1′′′, C-1′′′′), 171.2
C-8′′′, C-10′′′′), 135.7 (C-14′′′, C-16′′′′), 133.1 (C-9′′′, C-11′′′′),
2
-2′′′′), 2.24 (2H, m, Hb-12′′′, Hb-14′′′′), 2.12 (4H,
-17′′′′), 1.83 (2H, m, Ha-8′′′, Ha-8′′′′), 1.64 (4H,
-4′′′, H -5′′′, H -4′′′′, H
m, H
2
2
m, H
2
2
2
2
2
2
-
5
(
2
2
1
3
3 3
-16′′′, H -18′′′′); C NMR (150 MHz,
1
2
3
the procedure of Ashton.
(
Enzymatic Hydrolysis of 1. A solution of 1 (0.6 mg) and
Lipase type XI (0.72 units, Sigma) in the presence of Triton
X-100 (2.5 mg) in boric acid/borax buffer (0.63 mL, pH 7.7)
was stirred at 38 °C for 12 h. The reaction was quenched with
AcOH (0.1 mL), and then EtOH (2 mL) was added to the
reaction mixture. The solvent was removed under reduced
1
3
4
2
26.7 (C-13′′′, C-15′′′′), 51.4 (C-11′′′, C-13′′′′), 48.8 (C-7′′′, C-9′′′′),
6.0 (C-2′′′, C-2′′′′), 35.6 (C-6′′′, C-8′′′′), 31.6 (C-6′′′′), 31.3 (C-
′′′, C-4′′′′), 31.0 (C-7′′′′), 28.9 (C-5′′′, C-5′′′′), 26.8 (C-3′′′, C-3′′′′),
4.5 (C-12′′′, C-14′′′′), 22.8 (C-15′′′, C-17′′′′), and 15.2 (C-16′′′,
1
3
1
C-18′′′′); C and H NMR data for sugars and glycerol moieties,
+
see Table 1; ESIMS (pos.) m/z 959 (M + Na) , HRESIMS (pos.)
m/z 959.4971 (M + Na) , calcd for C51
pressure, and the residue was purified using a silica gel column
+
H
80
O17Na, 959.4979.
9
(
hexane/EtOAc, 1:1) to yield OPDA (7, 0.4 mg, 62%), which
2
2
Arabidopside D (2): colorless amorphous solid; [R]
67.2° (c 0.69, MeOH); IR (KBr) νmax 3424, 1736, 1714, and
D
was identified as OPDA methyl ester derived from the hydro-
lysate with trimethylsilyldiazomethane, using GC analysis.
+
10
-
1 1
1
631 cm ; H NMR (600 MHz, CD
3
OD) δ 7.96 (2H, dd, J )
Bioassay. Ten seeds of cress (Lepidium sativum L.) were
placed on a filter paper (No. 1, Toyo) moistened with 500 µL
of test solution containing 0.01% Triton X-100 (v/v) in a 2.7
cm Petri dish and kept for 40 h at 24 °C in the dark, after
which the lengths of their roots were measured. Seedlings
cultured on the solution containing 0.01% Triton X-100 was
used as controls. Data are represented as mean values with
standard errors of three experiments.
2
.1, 5.8 Hz, H-10′′′, H-10′′′′), 6.21 (2H, dd, J ) 2.1, 5.8 Hz,
H-11′′′, H-11′′′′), 5.45 (4H, m, H-15′′′, H-16′′′, H-15′′′′, H-16′′′′),
.09 (2H, m, H-9′′′, H-9′′′′), 2.54 (2H, m, H-13′′′, H-13′′′′), 2.50
2H, m, Ha-14′′′, Ha-14′′′′), 2.37 (4H, m, H -2′′′, H -2′′′′), 2.24
2H, m, Hb-14′′′, Hb-14′′′′), 2.13 (4H, m, H -17′′′, H -17′′′′), 1.83
2H, m, Ha-8′′′, Ha-8′′′′), 1.65 (4H, m, H -3′′′, H -3′′′′), 1.36 (16H,
-4′′′, H -5′′′, H -6′′′, H -7′′′, H -4′′′′, H -5′′′′, H -6′′′′, H
′′′′), 1.23 (2H, m, Hb-8′′′, Hb-8′′′′) and 1.02 (6H, t, J ) 7.5 Hz,
3
(
(
(
2
2
2
2
2
2
m, H
2
2
2
2
2
2
2
2
-
7
1
3
H
3
-18′′′, H
2′′′, C-12′′′′), 176.8 (C-1′′′, C-1′′′′), 171.2 (C-10′′′, C-10′′′′), 135.7
C-16′′′, C-16′′′′), 134.1 (C-11′′′, C-11′′′′), 126.9 (C-15′′′, C-15′′′′),
3 3
-18′′′′); C NMR (150 MHz, CD OD) δ 215.2 (C-
1
Acknowledgment. We thank Dr. S. Matsuyama (Univer-
sity of Tsukuba) for chiral GC analysis, Banyu Pharmaceutical
Co., Ltd. for measurement of ESIMS, and Daiso Co., Ltd. for
(
4
9.8 (C-13′′′, C-13′′′′), 44.8 (C-9′′′, C-9′′′′), 36.0 (C-2′′′, C-2′′′′),
5.6 (C-8′′′, C-8′′′′), 31.5 (C-6′′′, C-6′′′′), 31.3 (C-4′′′, C-4′′′′), 30.9
3
(
R)- and (S)-glycerol acetonides. This work was partly sup-
(
C-7′′′, C-7′′′′), 29.3 (C-5′′′, C-5′′′′), 26.8 (C-3′′′, C-3′′′′), 24.7 (C-
ported by a Grant-in-Aid for JSPS Research Fellowships for
Young Scientists.
1
4′′′, C-14′′′′), 22.3 (C-17′′′, C-17′′′′) and 15.3 (C-18′′′, C-18′′′′);
13
1
C and H NMR data for sugars and glycerol moieties, see
+
Table 1; ESIMS (pos.) m/z 987 (M + Na) , HRESIMS (pos.)
m/z 987.5285, calcd for C51
80
H O17Na, 987.5293.
References and Notes
Methanolysis of 2 and Chiral GC Analyses of the
Methanolysates. After a solution of 2 (0.1 mg) in 0.6 mol/L
HCl/MeOH (0.1 mL) was stirred at 60 °C for 45 min, the
solvent was removed under reduced pressure. The residue
was partitioned between hexane and 90% MeOH, and the
(1) Hisamatsu, Y.; Goto, N.; Hasegawa, K.; Shigemori, H. Tetrahedron
Lett. 2003, 44, 5553-5556.
(
2) Baertschi, S. W.; Ingram, C. D.; Harris, T. M.; Brash, A. R.
Biochemistry 1988, 27, 18-24.
(3) Weber, H.; Vick, B. A.; Farmer, E. E. Proc. Natl. Acad. Sci. U.S.A.
1997, 94, 10473-10478.
(
7
hexane-soluble materials were used for chiral GC analyses.
4) Hisamatsu, Y.; Hasegawa, K.; Shigemori, H. Recent Res. Dev. Org.
Chem. 2004, 8, 467-476.
5) Hansen, P. E. Prog. NMR Spectrosc. 1981, 14, 175-296.
6) Gunstone, F. D.; Pollard, M. R.; Scrimgeour, C. M.; Vedanayagam,
H. S. Chem. Phys. Lipids 1977, 18, 115-129.
Chiral GC conditions: column: γ-DEX 120 capillary column,
SUPELCO; program rate: 40 f 160 °C (at 10 °C/min), 160 °C
(
(
(
360 min), 160 f 200 °C (at 10 °C/min), and 200 °C (60 min).
The absolute configurations of the methyl ester of OPDA

Notes
Journal of Natural Products, 2005, Vol. 68, No. 4 603
(
(
(
7) Laudert, D.; Hennig, P.; Stelmach, B. A.; M u¨ ller, A.; Andert, L.;
(11) Stelmach, B. A.; M u¨ ller, A.; Hennig, P.; Gebhardt, S.; Schubert-
Zsilavecz M.; Weiler, E. W. J. Biol. Chem. 2001, 276, 12832-12838.
(12) Ashton, W. T.; Canning, L. F.; Reynolds, G. F.; Tolman, R. L.; Karkas,
J. D.; Linou, R.; Davies, M.-E. M.; Dewitt, C. M.; Perry, H. C.; Field,
A. K. J. Med. Chem. 1985, 28, 926-933.
Weiler, E. W. Anal. Biochem. 1997, 246, 211-217.
8) Uzawa, H.; Nishida, Y.; Ohrui, H.; Meguro, H. Agric. Biol. Chem.
1
989, 53, 2327-2333.
9) Murakami, N.; Morimoto, T.; Imamura, H.; Nagatsu, A.; Sakakibara,
J. Tetrahedron 1994, 50, 1993-2002.
(
10) Hashimoto, N.; Aoyama, T.; Shioiri, T. Chem. Pharm. Bull. 1981, 29,
1475-8.
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