Effect of the benzylic structure of lignan on antioxidant activity

Article abstract of DOI:10.1271/bbb.70275

The effect of the benzylic structure of lignan on antioxidant activity was evaluated. Secoisolariciresinol (1) and 3,4-bis(4-hydroxy-3-methoxybenzyl) tetrahydrofuran (2), which have two secondary benzylic positions without oxygen, showed the highest antioxidant activity. Optically active verrucosin (4) was synthesized for the first time in this experiment.

Full text of DOI:10.1271/bbb.70275

Biosci. Biotechnol. Biochem., 71 (9), 2283–2290, 2007
Effect of the Benzylic Structure of Lignan on Antioxidant Activity
y
Satoshi YAMAUCHI,1; Takuya SUGAHARA,1 Junko MATSUGI,1 Tatsushi SOMEYA,2
Toshiya MASUDA,2 Taro KISHIDA,1 Koichi AKIYAMA,3 and Masafumi MARUYAMA
1
1Faculty of Agriculture, Ehime University, 3-5-7 Tarumi, Matsuyama, Ehime 790-8566, Japan
2Faculty of Integrated Arts and Sciences, University of Tokushima, Tokushima 770-8502, Japan
3Integrated Center for Sciences, Ehime University, 3-5-7 Tarumi, Matsuyama, Ehime 790-8566, Japan
Received May 7, 2007; Accepted June 4, 2007; Online Publication, September 7, 2007
[doi:10.1271/bbb.70275]
The effect of the benzylic structure of lignan on
antioxidant activity was evaluated. Secoisolariciresinol
(1) and 3,4-bis(4-hydroxy-3-methoxybenzyl)tetrahydro-
furan (2), which have two secondary benzylic positions
without oxygen, showed the highest antioxidant activity.
Optically active verrucosin (4) was synthesized for the
first time in this experiment.
other is oxidized and contained in the 5-membered ring.
Both benzylic positions of verrucosin (4) are oxidized
and contained in the five-membered ring. Both benzylic
positions of pinoresinol (5) are also oxidized and con-
tained in the furofuran ring. A comparison of antioxidant
activity of 1–5 will clarify the effect of the benzylic
structure of lignan on the activity. Compounds 1–5 are
common lignans contained in plant food and natural
resources, and the effect of antioxidant activity on health
has been suggested.7) The result of this present study
might contribute to biological research and suggest the
effective utilization of natural resources containing
lignans.
Most of the previous studies have examined the
antioxidant activity of compounds isolated in one
experiment; however, this methodology is not sufficient
for research about the relationship between structure and
activity. In this present study, organic synthesis was
employed in collecting the required compounds to
compare the effect of the benzylic structure. Lignans
3–5 were synthesized by a new synthetic route. The
stereoselective synthesis of the tetra-substituted tetrahy-
drofuran lignan, verrucosin (4), is described in this
article.
Key words: lignan; antioxidant activity; benzylic struc-
ture
The antioxidant activity of phenolic lignan is known,
as well as many kinds of biological activity. Although
the importance of the phenolic group for antioxidant
activity is well known, the effect of the structure, except
for the aryl group, on this antioxidant activity has not
been elucidated. Some research has been done1–5) to
clarify the relationship between the structure and anti-
oxidant activity of lignan. This research showed that
lignans bearing a lower oxidation degree at the benzylic
position had higher activity3,4) and that the presence of a
tertiary hydroxy group decreased this activity.1,2) Eklund
and co-workers have shown the production of a benzylic
oxidized product by treating matairesinol with 2,2-
diphenyl-1-picrylhydrazyl (DPPH)5) or 2,3-dichloro-5,6-
dicyano-1,4-benzoquinone (DDQ).6) These studies have
suggested the important role of the benzylic position in
antioxidant activity; however, only the oxidation degree
at the benzylic position has been compared. Each lignan
has particular structural characteristics at the benzylic
position. Studies at the effect of the benzylic structure on
antioxidant activity would lead to an evaluation of the
antioxidant activity of lignans. To define the relationship
between the benzylic structure and antioxidant activity,
five common lignans (1–5) having differences in their
benzylic structure were selected (Fig. 1). Secoisolaricir-
esinol (1) and 3,4-bis(4-hydroxy-3-methoxybenzyl)tetra-
hydrofuran (2) have two secondary benzylic positions
without oxygen. One of the benzylic positions of
lariciresinol (3) is secondary without oxygen, and the
Results and Discussion
Synthesis of lignans
The syntheses of 1–5 are shown in the Scheme 1.
Secoisolariciresinol (1)8) was synthesized from aldol
product 63) by LiBH4 reduction and desilylation with
tetra-n-butylammonium fluoride, this being followed by
a treatment with H2 in the presence of Pd0. 3,4-Bis(4-
hydroxy-3-methoxybenzyl)tetrahydrofuran (2) was syn-
thesized by the previously reported method.3) Laricir-
esinol (3)9) was converted from butanediol 7.3) After
desilylating with using tetra-n-butylammonium fluoride,
resulting triol 8 was treated with a catalytic amount of
10-camphorsulfonic acid, giving SN1 intramolecular
cyclization product 9. Hydrogenolysis of 9 gave laricir-
y
To whom correspondence should be addressed. Fax: +81-89-977-4364; E-mail: syamauch@agr.ehime-u.ac.jp

2284
S. YAMAUCHI et al.
Ar2
H
O
Ar2
H
Ar2
Ar2
O
O
Ar2
Ar2
O
Ar2
Ar2
H
H
OH
OH
O
Ar2
Ar2
HO
1
2
3
4
5
Fig. 1. Common Lignans.
Ar2, 4-hydroxy-3-methoxyphenyl
TIPSO
Ar1
OH
H
H
O
7
Ar1
HO
OH
HO
OH
9'
8'
Ar1
b
ref.
c
a
8
3
H
H
H
H
H
H
1
9
TIPSO
OH
Ar1
Ar1
HO
Ar1
O
O
Ar1
Ar1
H
H
6
8
9
7
H
TIPSO
Ar1
H
7
O
7'
Ar1
Ar1
Ar1
Ar1
O
O
7
d
e
g
PivO
6
H
H
4
8'
9'
Ar1
8
H
9
R
PivO
OPiv
9'
10
12: R = OH
13: R = H
OPiv
f
11
HO
Ar1
TIPSO
Ar1
Ar1
O
H
OMOM
H
H
OMOM
H
Ar1
7'
OH
H
HO
9
9
8
Ar1
Ar1
7'
h
i
j
ref.
H
H
H
H
H
H
H
8
5
8'
9'
8'
9'
Ar1
OH
O
7
O
Ar1
7
O
O
HO
14
15
16
17
Scheme 1. Synthesis of Lignans.
Ar1, 4-benzyloxy-3-methoxyphenyl. (a) (1) LiBH4, THF, 0 ꢀC, 16 h; (2) n-Bu4NF, THF, r.t., 7 h (74% yield, 2 steps); (3) H2, Pd0, r.t., 3 h (64%
yield); (b) n-Bu4NF, THF, r.t., 1 h (97% yield); (c) CSA, CH2Cl2, r.t., 16 h (61% yield); (d) (1) LiBH4, THF, 0 ꢀC, 16 h; (2) PivCl, pyridine, r.t.,
2 h; (3) PCC, MS 4A, CH2Cl2, r.t., 16 h (48% yield, 3 steps); (e) (1) n-Bu4NF, AcOH, THF, r.t., 1 h; (2) H2, Pd(OH)2, EtOAc, r.t., 15 min; (3)
BnBr, K2CO3, dibenzo-18-C-6, CH3CN, reflux, 16 h; (4) 1 M aq. NaOH, EtOH, r.t., 16 h (29%, 4 steps); (f) (1) TsCl, pyridine, r.t., 2 h (78%); (2)
LiAlH4, THF, r.t., 30 min (50% yield); (g) H2, 5%Pd/C, EtOAc, r.t., 5 h (58%); (h) (1) MOMCl, iso-Pr2NEt, CH2Cl2, r.t., 16 h; (2) DIBAL-H,
toluene, ꢁ70 ꢀC, 30 min; (3) NaBH4, EtOH, r.t., 1 h; (4) n-Bu4NF, THF, r.t., 1 h (77% yield, 4 steps); (i) CSA, CH2Cl2, 0 ꢀC, 18 h (82% yield); (j)
6 M aq. HCl, THF, r.t., 4 h (77% yield).
esinol (3). Verrucosin (4) was obtained from aldol pro-
duct 6.3) Aldol product 6 was converted to benzyl ketone
10 by the previously described method.10) Desilylation
of 10 in a tetra-n-butylammonium fluoride-acetic acid
system and a subsequent treatment with H2 in the
presence of Pd(OH)2 gave 12 as a single isomer. The
reaction mechanism could be assumed to involve the
hemiacetal formed by desilylation being transformed to
dihydrofuran 11. The reduction of the newly produced
allylic 90 position of 11 easily occurred, and hydro-
genation of 11 was then performed from the opposite
side of the aryl group at the 7 position.11) Selective
reduction of one of the pivaloyloxymethyl groups to a
methyl group was observed in this experiment. Treat-
ment of 12 with tosyl chloride gave a tosylate which was
then subjected to LiAlH4 reduction to give compound
13. Hydrogenolysis of 13 by Pd/C under H2 gas gave
verrucosin (4).12) The stereoselective synthesis of verru-
cosin (4) was achieved for the first time. Pinoresinol (5)
was synthesized from 14.3) After aldol product 14 had
been transformed to tetrahydrofuran derivative 16 by the
previously described method,13) cyclization to the
furofuran ring by using an aqueous HCl solution was
performed, giving furofuran compound 17.9) Hydro-
genolysis gave pinoresinol (5).
Antioxidant activity of 1–5
To evaluate the antioxidant activity, ethyl linoleate
was treated with compounds 1–5 and 2,2-diphenyl-1-
picrylhydrazyl (DPPH), and then the concentration of
ethyl linoleate hydroperoxide produced was calculated
(Fig. 2). The results showed a different activity level
between compounds 1 and 2 bearing a secondary ben-
zylic position without oxygen and compounds 4 and 5
bearing two oxidized benzylic positions enclosed in a
ring. It could be assumed that the presence of the

Antioxidant Activity of Lignan
2285
Control
Trolox
Ferulic acid
1
2
3
4
5
0
2
4
6
8
10
12
Ethyl linoleate hydroperoxide (mM)
Fig. 2. Inhibition of the Production of the Ethyl Linoleate Hydroperoxide of Lignans 1–5.
APPH, 10 mM; sample, 0.05 mM; ferulic acid, 0.05 mM; Trolox, 0.05 mM; ethyl linoleate, 50 mM.
Sesamol
1
2
3
4
5
0
10
20
30
40
50
60
70
80
Relative radical scavenging activity ( % )
Fig. 3. Radical Scavenging Activity of Lignans 1–5.
Sample, 20 mM (MeOH); DPPH, 0.1 mM (MeOH); reaction, 30 min at r.t., n ¼ 3.
secondary benzylic position without oxygen was im-
portant for inhibiting ethyl linoleate hydroperoxide
production. The activity of 1 and 2 was higher than
that of the known naturally occurring antioxidant com-
ponent, ferulic acid. The activity level of 3–5 was almost
same as that of ferulic acid. The different level of anti-
oxidant activity of 1–5 was clearer in their radical
scavenging activity (Fig. 3). The activity of dibenzyl
compounds 1 and 2 was higher than that of 3 which has
one secondary benzylic position without oxygen and one
oxidized benzylic position in the tetrahydrofuran ring.
The activity of compound 3 was higher than that of com-
pounds 4 and 5 bearing two oxidized benzylic positions
in the ring. The lignans bearing a secondary benzylic
position without oxygen showed stronger activity. The
oxygen-free secondary benzylic position would play an
important role in the stability of the phenoxy radical.
The activity of acyclic lignan 1 was higher than that of
cyclic lignan 2, showing almost same activity level as
that of sesamol. The two phenolic groups of 2,3-di-
benzyltetrahydrofuran-type lignan 2 was closer than that
of acyclic lignan 1. It could be assumed that the two
phenoxy radicals of 1 were more stable than the closer
phenoxy radicals of 2, and that the radical scavenging
activity of 1 was higher than that of 2. By comparing the
activity of 4 with 5, it could be assumed that the lignan
bearing closer phenolic groups had weaker radical
scavenging activity. The superoxide dismutase-like
activity of compounds 1–5 was also examined (Fig. 4).
The compounds bearing secondary benzylic positions

2286
S. YAMAUCHI et al.
Sesamol
1
2
3
4
5
0
2
4
6
8
10
SOD activity ( unit/ml )
Fig. 4. SOD-Like Activity of 1–5.
Sample, 160 mM; assay, DOS assay kitWET from Dojin; reaction, r.t., 90 min, 37 ꢀC.
without oxygen (1 and 2) showed higher activity than
the other compounds. It was thus found that the oxygen-
free secondary benzylic position was also important for
superoxide dismutase-like activity.
The antioxidant activity of five common lignans
having differences in the benzylic structure was exam-
ined in this experiment. It was found that the lignans
bearing an oxygen-free secondary benzylic position had
higher antioxidant activity. The relationship between the
benzylic structure of common lignans and their anti-
oxidant activity is shown for the first time, and it is
suggested that the antioxidant level of a lignan depends
on the benzylic structure.
solution was separated washed with sat. aq. CuSO4
solution, sat. aq. NaHCO3 solution, and brine, and dried
(Na2SO4). Concentration followed by silica gel column
chromatography (EtOAc/hexane = 6/1) gave a tetraol
20
(1.27 g, 2.21 mmol, 74%) as a colorless oil, ½ꢀꢂ
¼
D
þ18 (c 1.1, CHCl3). NMR ꢁH (CDCl3): 2.00 (1H, m,
CH), 2.08 (1H, m, CH), 3.65 (3H, s, OCH3), 3.66–3.86
(4H, m, CH2OH), 3.74 (3H, s, OCH3), 4.05–4.70 (4H,
br., OH), 4.49 (1H, br. s, ArCHOH), 4.83 (1H, br. s,
ArCHOH), 5.03 (2H, s, ArCH2O), 5.06 (2H, s,
ArCH2O), 6.45–6.48 (2H, m, ArH), 6.63 (1H, d, J
8.3 Hz, ArH), 6.68 (1H, d, J 8.2 Hz, ArH), 6.73 (1H, s,
ArH), 6.78 (1H, d, J 8.3 Hz, ArH), 7.25–7.32 (6H, m,
ArH), 7.34–7.40 (4H, m, ArH). NMR ꢁC (CDCl3): 44.4,
49.0, 55.8, 56.0, 58.5, 61.8, 70.96, 70.99, 73.9, 76.4,
109.5, 109.7, 113.5, 113.7, 117.9, 118.1, 127.3, 127.8,
127.9, 128.48, 128.51, 136.0, 136.3, 137.0, 147.1, 147.3,
149.5. FABMS m=z (%): 575 (ðM þ HÞþ, 1), 307 (42),
154 (100), 136 (70). HRFABMS ðM þ HÞþ: Calcd. for
C34H39O8, 575.2645. Found, 575.2642. A mixture of
PdCl2 (0.4 g) in EtOH (10 ml) was stirred under H2 gas
at ambient temperature for 5 min. After removal of
EtOH by decantation, a solution of the tetraol (1.27 g,
2.21 mmol) in THF (20 ml) was added, and then the
reaction mixture was stirred under H2 gas at ambient
temperature for 5 h before filtration. After concentration
of the filtrate, the residue was applied to silica gel
column chromatography (EtOAc/hexane = 6/1) to give
(+)-secoisolariciresinol (1) (0.51 g, 1.41 mmol, 64%) as
Experimental
Melting point (mp) data are uncorrected. NMR data
were measured by a JNM-EX400 spectrometer, using
TMS as a standard (0 ppm), MS data were measured
with a JMS-MS700V spectrometer, and optical rotation
values were evaluated with a Horiba SEPA-200 instru-
ment. The silica gel used was Wakogel C-300 (Wako,
200–300 mesh). The numbering of compounds follows
IUPAC nomenclatural rules.
(+)-Secoisolariciresinol (1). To an ice-cooled sus-
pension of LiBH4 (0.51 g, 23.4 mmol) in THF (20 ml)
was added a solution of erythro aldol product 6 (2.18 g,
3.00 mmol) in THF (20 ml), and then the reaction
mixture was stirred at 0 ꢀC for 16 h before addition of
sat. aq. NH4Cl solution. After concentration, the residue
was dissolved in EtOAc and H2O. The organic solution
was separated, washed with brine, and dried (Na2SO4).
Concentration gave a crude triol. To an ice-cooled
solution of this crude triol in THF (15 ml) was added n-
Bu4NF (3.70 ml, 1 M in THF, 3.70 mmol), and then the
reaction solution was stirred at room temperature for 7 h
before addition of sat. aq. NH4Cl solution. The organic
colorless crystals, mp 111–112 ꢀC (CHCl3-(iso-Pr)2O),
20
114–115 ꢀC in literature,8) ½ꢀꢂ
¼ þ29 (c 0.3, ace-
D
tone). The NMR data agreed with those in the liter-
ature.8)
(1R,2S,3S)-3-(4-Benzyloxy-3-methoxybenzyl)-1-(4-ben-
zyloxy-3-methoxyphenyl)-2-hydroxymethyl-1,4-butanedi-
ol (8). A reaction solution of silyl ether 73) (0.65 g,
0.91 mmol) and n-Bu4NF (1.00 ml, 1 M THF, 1.00 mmol)

Antioxidant Activity of Lignan
2287
in THF (20 ml) was stirred at room temperature for 1 h
before addition of sat. aq. NH4Cl solution. The organic
solution was separated, washed with sat. aq. CuSO4
solution, sat. aq. NaHCO3 solution, and brine, and then
dried (Na2SO4). Concentration followed by silica gel
column chromatography (EtOAc/hexane = 3/2 and
(0.90 ml, 7.31 mmol). After the reaction mixture was
stirred at room temperature for 2 h, EtOAc and H2O
were added. The organic solution was separated, washed
with 6 M aq. HCl solution, sat. aq. NaHCO3 solution, and
brine, and then dried (Na2SO4). Concentration gave a
crude diPiv ester. A reaction mixture of diPiv ester, PCC
(0.55 g, 2.55 mmol), and MS 4A (0.1 g) in CH2Cl2 (20
ml) was stirred at room temperature for 16 h before
addition of dry ether. After filtration, the filtrate was
concentrated. The residue was applied to silica gel
column chromatography (EtOAc/hexane = 1/4) to
9/1) gave triol 8 (0.49 g, 0.88 mmol, 97%) as a colorless
20
oil, ½ꢀꢂ
¼ þ33 (c 0.8, CHCl3). NMR ꢁH (CDCl3):
D
1.89 (1H, br. s, OH), 2.16 (1H, m, CH), 2.30–2.38 (2H,
m, CH, OH), 2.51 (1H, dd, J 13.8, 8.2 Hz, ArCHH), 2.59
(1H, dd, J 13.8, 7.3 Hz, ArCHH), 3.46 (1H, dd, J 9.8,
8.9 Hz, CHHOH), 3.59–3.70 (2H, m, CH2OH), 3.68
(3H, s, OCH3), 3.74 (3H, s, OCH3), 3.89 (1H, m,
CHHOH), 4.80 (1H, br. s, OH), 4.94 (1H, br. dd, J 5.9,
5.9 Hz, ArCHOH), 5.01 (2H, s, OCH2Ar), 5.06 (2H, s,
OCH2Ar), 6.45 (1H, d, J 9.4 Hz, ArH), 6.57 (1H, d, J
8.2 Hz, ArH), 6.61–6.74 (3H, m, ArH), 6.91 (1H, s,
ArH), 7.25–7.36 (6H, m, ArH), 7.39–7.41 (4H, m, ArH).
NMR ꢁC (CDCl3): 36.3, 38.0, 48.6, 55.8, 60.5, 61.7,
70.9, 72.7, 109.4, 113.5, 114.1, 114.3, 117.5, 117.8,
127.2, 127.4, 128.4, 128.5, 131.7, 136.2, 136.4, 137.1,
146.9, 147.1, 148.7, 149.2. EIMS m=z: 558 (Mþ, 6), 91
(100). HREIMS m=z: 558.2617 (calcd. for C34H38O7,
558.2617).
give ketone 10 (1.49 g, 1.66 mmol, 48%, 3 steps) as a
20
colorless oil, ½ꢀꢂ
¼ ꢁ3 (c 1, CHCl3). NMR ꢁH
D
(CDCl3): 0.08–1.03 (21H, m, iso-Pr), 1.00 (9H, s, tert-
Bu), 1.17 (9H, s, tert-Bu), 2.59 (1H, m, 3-H), 3.72 (3H,
s, OCH3), 3.90 (3H, s, OCH3), 4.04 (1H, dd, J 11.8,
5.6 Hz, CHHOPiv), 4.11–4.17 (1H, m, 2-H), 4.17 (1H,
dd, J 11.8, 4.2 Hz, CHHOPiv), 4.41 (1H, dd, J 10.7,
9.3 Hz, CHHOPiv), 4.50 (1H, dd, J 10.7, 9.3 Hz,
CHHOPiv), 4.90 (1H, d, J 5.9 Hz, ArCHOSi), 5.10
(2H, s, OCH2Ph), 5.22 (2H, s, OCH2OPh), 6.56 (1H, d, J
6.3 Hz, ArH), 6.71 (1H, d, J 7.8 Hz, ArH), 6.81–6.83
(4H, m, ArH), 7.26–7.44 (9H, m, ArH), 7.52 (1H, d, J
2.0 Hz, ArH). NMR ꢁC (CDCl3): 12.5, 18.0, 18.1, 27.0,
27.1, 38.5, 38.7, 43.5, 46.5, 55.7, 56.0, 61.5, 63.1, 70.8,
71.1, 73.6, 110.6, 111.2, 111.3, 112.0, 113.7, 119.1,
123.0, 127.2, 127.8, 128.1, 128.5, 128.7, 130.0, 135.4,
136.2, 137.0, 147.7, 149.6, 149.7, 152.6, 177.8, 178.0,
198.3. Anal. Found: C, 71.13; H, 8.16. Calcd. for
C53H72O10Si: C, 70.95; H, 8.09%.
(7R,8S,80S)-4,40-Dibenzyloxy-3,30-dimethoxy-7,90-epox-
ylignan-9-ol (9). A reaction solution of triol 8 (0.10 g,
0.18 mmol) and 10-camphorsulfonic acid (5 mg, 0.022
mmol) in CH2Cl2 (5 ml) was stirred at room temperature
for 16 h before addition of Et3N. After concentration, the
residue was applied to silica gel column chromatography
(EtOAc/hexane = 1/2 and 2.5% MeOH in CHCl3) to
(7S,70R,8R,80S)-4,40-Dibenzyloxy-3,30-dimethoxy-7,70-
epoxylignan-90-ol (12). A reaction solution of silyl ether
10 (1.49 g, 1.67 mmol), n-Bu4NF (4.80 ml, 1 M in THF,
4.80 mmol), and AcOH (0.15 ml) in THF (20 ml) was
stirred at room temperature for 1 h before addition of sat.
aq. NaHCO3 solution and EtOAc. The organic solution
was separated, washed with sat. aq. CuSO4 solution,
sat. aq. NaHCO3 solution, and brine, and then dried
(Na2SO4). Concentration gave an unstable hemiacetal.
A reaction mixture of this crude hemiacetal and 20%
Pd(OH)2/C (0.70 g) in EtOAc (10 ml) was stirred under
H2 gas at ambient temperature for 15 min. After
filtration, the filtrate was concentrated to give a crude
tetrahydrofuran derivative bearing a phenol. A reaction
mixture of this crude tetrahydrofuran derivative bearing
a phenol, K2CO3 (1.17 g, 8.47 mmol), benzyl bromide
(0.40 ml, 3.36 mmol), and dibenzo-18-C-6 (10 mg, 0.028
mmol) in CH3CN (40 ml) was heated under refluxing for
16 h before concentration. The residue was dissolved in
H2O and EtOAc. The organic solution was separated,
washed with brine, and dried (Na2SO4). After concen-
tration, the residue was dissolved in EtOH (12 ml) and
1 M aq. NaOH solution (8 ml), and then the reaction
solution was stirred at room temperature for 16 h before
additions of CHCl3 and H2O. The organic solution was
separated, washed with brine, and dried (Na2SO4).
Concentration followed by silica gel column chroma-
give tetrahydrofuran derivative 9 (57 mg, 0.11 mmol,
20
61%) as a colorless oil, ½ꢀꢂ
¼ ꢁ9 (c 0.7, CHCl3). The
D
NMR data agreed with those in the literature.9) EIMS
m=z: 540 (Mþ, 17), 91 (100). HREIMS m=z: 540.2513
(calcd. for C34H36O6, 540.2512).
(7R,8S,80S)-3,30-Dimethoxy-7,90-epoxylignane-4,40,9-
triol ((ꢁ)-lariciresinol) (3). Hydrogenolysis of 9 gave
20
(ꢁ)-lariciresinol, mp 156–158 ꢀC, ½ꢀꢂ
¼ ꢁ16 (c 0.7,
D
25
CHCl3), mp 162–164 ꢀC, ½ꢀꢂ
¼ ꢁ17:8 (c 1.4, ace-
D
tone) in the literature,14) The NMR data agreed with
those in the literature.9) EIMS m=z: 360 (Mþ, 100), 137
(67). HREIMS m=z: 360.1573 (calcd. for C20H24O6,
360.1573).
(2S,3S,4R)-1,4-Bis(4-benzyloxy-3-methoxyphenyl)-2,3-
bis(pivaloyloxymethyl)-4-triisopropylsilyloxy-1-butanone
(10). To an ice-cooled solution of LiBH4 (0.62 g, 28.5
mmol) in THF (10 ml) was added a solution of aldol
product 63) (2.54 g, 3.49 mmol) in THF (10 ml). The
reaction mixture was stood at 0 ꢀC for 16 h before
addition of sat. aq. NH4Cl solution. After concentration,
the residue was dissolved in EtOAc and H2O. The
organic solution was separated, washed with brine, dried
(Na2SO4), and concentrated. To an ice-cooled solution
of the residue in pyridine (10 ml) was added PivCl

2288
S. YAMAUCHI et al.
tography (EtOAc/hexane = 1/2, 1/1, and 3/1) gave
alcohol 12 (0.26 g, 0.48 mmol, 29%, 4 steps) as a col-
Found: C, 78.02; H, 6.81. Calcd. for C34H36O5: C,
77.84; H, 6.92%.
20
orless oil, ½ꢀꢂ
¼ ꢁ16 (c 1.3, CHCl3). NMR ꢁH
D
(CDCl3): 0.70 (3H, d, J 6.8 Hz, 9-H), 1.45 (1H, m, OH),
1.99 (1H, m, 8-H), 2.46 (1H, m, 80-H), 3.77–3.80 (2H,
m, CH2OH), 3.86 (3H, s, OCH3), 3.90 (3H, s, OCH3),
4.68 (1H, d, J 7.8 Hz, 70-H), 5.06 (1H, d, J 6.8 Hz, 7-H),
5.14 (2H, s, OCH2Ph), 5.15 (2H, s, OCH2Ph), 6.82–6.91
(4H, m, ArH), 6.98 (1H, dd, J 8.3, 2.0 Hz, ArH), 7.10
(1H, d, J 2.0 Hz, ArH), 7.27–7.38 (6H, m, ArH), 7.43–
7.44 (4H, m, ArH). NMR ꢁC (CDCl3): 16.6, 40.8, 56.0,
57.0, 63.0, 71.1, 71.2, 82.6, 82.8, 110.5, 110.6, 113.9,
114.0, 118.6, 118.7, 127.2, 127.3, 127.7, 127.8, 128.45,
128.50, 133.2, 134.8, 137.2, 137.3, 147.2, 147.8, 149.4,
149.8. Anal. Found: C, 75.79; H, 6.79. Calcd. for
C34H36O6: C, 75.53; H, 6.71%.
(7S,70R,8R,80R)-3,30-Dimethoxy-7,70-epoxylignane-4,40-
diol (verrucosin) (4). A reaction mixture of benzyl ether
13 (34 mg, 0.065 mmol) and 5% Pd/C (60 mg) in EtOAc
(15 ml) was stirred under H2 gas at ambient temperature
for 5 h before filtration. The filtrate was concentrated,
and then the residue was applied to silica gel column
chromatography (EtOAc/hexane = 1/1) to give verru-
cosin (4) (13 mg, 0.038 mmol, 58%) as a colorless oil,
20
21
½ꢀꢂ D ꢁ48 (c 0.3 CHCl3), ½ꢀꢂ D ꢁ30:0 (c 0.90, CHCl3)
in the literature.15) The NMR data agreed with those in
the literature.12) EIMS m=z: 344 (Mþ, 67), 192 (100),
177 (80), 145 (75). HREIMS m=z: 344.1624 (calcd. for
C20H24O5, 344.1624).
(7S,70R,8R,80R)-4,40-Dibenzyloxy-3,30-dimethoxy-7,70-
epoxylignane (13). A reaction solution of alcohol 12
(0.32 g, 0.59 mmol) and TsCl (0.34 g, 1.78 mmol) in
pyridine (1 ml) was stirred at room temperature for 2 h
before additions of H2O and EtOAc. The organic
solution was separated, washed with 6 M aq. HCl
solution, sat. aq. NaHCO3 solution, and brine, and then
dried (Na2SO4). Concentration followed by silica gel
column chromatography (EtOAc/hexane = 1/3) gave
an unstable tosylate (0.32 g, 0.46 mmol, 78%) as a col-
orless oil, NMR ꢁH (CDCl3): 0.63 (3H, d, J 7.0 Hz,
CH3), 2.00 (1H, m), 2.43 (3H, s, p-CH3Ph), 2.43 (1H,
m), 3.86 (3H, s, OCH3), 3.87 (3H, s, OCH3), 4.15 (2H,
d, J 5.2 Hz, CH2OTs), 4.60 (1H, d, J 8.2 Hz, ArCH-
OCH2), 5.01 (1H, d, J 7.3 Hz, ArCHOCH2), 5.13 (2H, s,
OCH2Ar), 5.15 (2H, s, OCH2Ar), 6.77–6.82 (3H, m,
ArH), 6.85–6.87 (2H, m, ArH), 6.98 (1H, d, J 1.5 Hz,
ArH), 7.27–7.38 (8H, m, ArH), 7.42–7.45 (4H, m, ArH),
7.76 (2H, d, J 8.3 Hz, ArH). To an ice-cooled suspension
of LiAlH4 (0.22 g, 5.80 mmol) in THF (10 ml) was
added a solution of this tosylate (0.32 g, 0.46 mmol) in
THF (10 ml). The resulting reaction mixture was stirred
at room temperature for 2 h before additions of sat. aq.
MgSO4 solution and K2CO3. The resulting mixture was
stirred at room temperature for 30 min and then filtered.
The filtrate was concentrated, and the resulting residue
was applied to silica gel column chromatography
(1R,2R,3R)-1-(4-Benzyloxy-3-methoxyphenyl)-3-[(S)-
(4-benzyloxy-3-methoxyphenyl)(methoxymethoxy)meth-
yl]-2-hydroxymethyl-1,4-butanediol (15). To a solution
of aldol product 143) (1.34 g, 1.84 mmol) and iso-Pr2NEt
(15.4 ml, 88.4 mmol) in CH2Cl2 (20 ml) was added
MOMCl (3.40 ml, 44.8 mmol). The resulting reaction
mixture was stirred at room temperature for 16 h before
additions of MeOH (14 ml) and CH2Cl2. The organic
solution was washed with 1 M aq. HCl solution, sat. aq.
NaHCO3 solution, and brine, and then dried (Na2SO4).
Concentration gave a crude MOM ether. To a solution of
this crude MOM ether in toluene (25 ml) was added
DIBAL-H (3.60 ml, 1 M in toluene, 3.60 mmol) at
ꢁ70 ꢀC. The reaction solution was stirred at ꢁ70 ꢀC
for 30 min, and then aq. 1 M HCl solution and EtOAc
were added. The organic solution was separated, washed
with sat. aq. NaHCO3 solution, and brine, and then died
(Na2SO4). After concentration, the residue was dis-
solved in EtOH. To the ice-cooled solution of the EtOH
solution was added NaBH4 (1.00 g, 26.4 mmol). After
the reaction mixture was stirred at room temperature for
1 h, 1 M aq. HCl solution was added. The mixture was
neutralized with sat. aq. NaHCO3 solution before
concentration. The resulting residue was dissolved in
H2O and EtOAc. The organic solution was separated,
washed with brine, and dried (Na2SO4). Concentration
gave a crude diol. A reaction solution of this crude diol
and n-Bu4NF (1.90 ml, 1 M in THF, 1.90 mmol) in THF
(20 ml) was stirred at room temperature for 1 h before
addition of sat. aq. NH4Cl solution. The organic solution
was separated, washed with sat. aq. CuSO4 solution,
sat. aq. NaHCO3 solution, and brine, and then dried
(Na2SO4). Concentration followed by silica gel column
chromatography (EtOAc/hexane = 1/1 and 3/1) gave
(hexane/EtOAc = 3/1) to give dimethyl compound 13
20
(0.12 g, 0.23 mmol, 50%) as a colorless oil, ½ꢀꢂ
¼
D
ꢁ27 (c 0.6, CHCl3). NMR ꢁH (CDCl3): 0.65 (3H, d, J
7.0 Hz, 90-H), 1.06 (3H, d, J 6.5 Hz, 9-H), 1.78 (1H, m,
8-H), 2.23 (1H, m, 80-H), 3.83 (3H, s, OCH3), 3.90 (3H,
s, OCH3), 4.40 (1H, d, J 9.3 Hz, 70-H), 5.11 (1H, d, J
8.6 Hz, 7-H), 5.14 (2H, s, OCH2Ph), 5.16 (2H, s,
OCH2Ph), 6.79–6.90 (4H, m, ArH), 6.96 (1H, dd, J 8.2,
1.9 Hz, ArH), 7.08 (1H, d, J 1.9 Hz, ArH), 7.29–7.38
(6H, m, ArH), 7.42–7.45 (4H, m, ArH). NMR ꢁC
(CDCl3): 15.0, 15.1, 46.0, 47.8, 55.97, 56.04, 71.15,
71.18, 83.1, 87.3, 110.7, 111.1, 113.8, 114.1, 118.6,
119.2, 127.26, 127.31, 127.75, 127.78, 128.47, 128.51,
134.1, 134.5, 137.28, 137.31, 147.8, 149.4, 149.7. Anal.
triol 15 (0.88 g, 1.42 mmol, 77%, 4 steps) as a colorless
20
oil, ½ꢀꢂ
¼ ꢁ47 (c 1.6, CHCl3). NMR ꢁH (CDCl3):
D
0.90 (1H, br. s, OH), 1.89 (1H, m, CH), 2.37 (1H, m,
CH), 3.31 (3H, s, OCH3), 3.48 (1H, dd, J 11.1, 5.5 Hz,
CHHOH), 3.53–3.68 (2H, br., OH), 3.74 (1H, dd, J 9.1,
2.1 Hz, CHHOH), 3.78 (3H, s, OCH3), 3.79 (3H, s,
OCH3), 3.91 (1H, br. d, J 9.1 Hz, CHHOH), 4.01 (1H,

Antioxidant Activity of Lignan
2289
dd, J 11.1, 4.8 Hz, CHHOH), 4.46 (2H, s, OCH2OCH3),
4.75 (2H, d, J 6.9 Hz, ArCHOMOM, ArCHOH), 5.11
(4H, s, OCH2Ph), 6.66–6.68 (3H, m, ArH), 6.73–6.80
(3H, m, ArH), 7.27–7.37 (6H, m, ArH), 7.41–7.43 (4H,
m, ArH). NMR ꢁC (CDCl3): 44.2, 47.7, 55.85, 55.89,
56.1, 59.6, 60.6, 70.9, 76.5, 77.8, 94.5, 109.9, 110.4,
113.4, 113.6, 118.7, 119.5, 127.2, 127.79, 127.82,
128.47, 128.49, 132.8, 136.2, 137.0, 137.1, 147.4,
147.6, 149.56, 149.59. HREIMS m=z: 618.2831 (calcd.
for C36H42O9, 618.2829).
agreed with those in the literature.9) EIMS m=z: 358
(Mþ, 75), 151 (100). HREIMS m=z: 358.1416 (calcd. for
C20H22O6, 358.1417).
Antioxidant activity of compounds 1–5 in a Tween 20
micelle system. The method of Masuda et al.17) was
slightly modified. To 160 ml of a DMSO solution of a
test sample (5.0 mM) were added freshly purified ethyl
linoleate (139 ml) and 0.3 M Tween 20–0.05 M phosphate
buffer (pH 7.4, 8 ml). The mixture was stirred vigo-
rously with a vortex mixer for 2 min and then treated in a
bath sonicator (Branson 2210) for 3 min to give a clear
micelle solution. Two ml of this micelle solution was put
into a straight vial (35 mm diameter; 75 mm height), and
100 ml of a 0.2 M AAPH aqueous solution was added to
the solution. After stirring again with the vortex mixer,
the vial was incubated at 37 ꢀC in the dark while
continuously shaking (82 shakes/min; Taitec P-11 water
bath shaker). After 3 h of incubation, a 20-ml aliquot
was taken from the solution and poured into 380 ml of a
methanolic solution of Trolox (0.2 mM). Ten ml of the
diluted solution was injected into the HPLC instrument
to analyze ethyl linoleate hydroperoxide under the fol-
lowing conditions: column, YMC-Pack ODS-A (4:6 ꢃ
150 mm); solvent, CH3CN–H2O = 9:1; flow rate, 1.0
ml/min; detection, 234 nm. The concentration of the
hydroperoxide was calculated from the peak area ob-
tained by the equation; Y ¼ 2:29X ꢃ 10ꢁ6 ꢁ 4:38 ꢃ
10ꢁ4, where Y is the concentration of ethyl linoleate
hydroperoxide (mM) and X is the peak area of hydro-
peroxide.
(7S,70S,8R,80R)-4,40-Dibenzyloxy-3,30-dimethoxy-70-
methoxymethoxy-7,90-epoxylignan-9-ol (16). A reaction
solution of triol 15 (0.88 g, 1.42 mmol) and 10-cam-
phorsulfonic acid (10 mg, 0.043 mmol) in CH2Cl2 (20
ml) was stood at 0 ꢀC for 18 h. After addition of a few
drops of Et3N, the mixture was concentrated. The res-
idue was applied to silica gel column chromatography
(EtOAc/hexane = 1/2 and 1/1) to give tetrahydrofuran
derivative 16 (0.70 g, 1.17 mmol, 82%) as colorless
20
crystals, mp 90–91 ꢀC (EtOH), ½ꢀꢂ
¼ ꢁ49 (c 0.9,
D
CHCl3). A triol (0.10 g, 0.16 mmol, 11%) was recov-
ered. NMR ꢁH (CDCl3): 2.56 (1H, m, CH), 2.88 (1H, m,
CH), 3.23 (1H, br. s, OH), 3.35–3.42 (1H, overlapped, 9-
HH), 3.40 (3H, s, OCH3), 3.68 (1H, dd, J 9.0, 6.4 Hz, 90-
HH), 3.77 (1H, m, 9-HH), 3.88 (6H, s, OCH3), 3.99 (1H,
dd, J 9.0, 9.0 Hz, 90-HH), 4.46 (2H, s, OCH2OCH3), 4.73
(1H, d, J 10.1 Hz, 70-H), 4.74 (1H, d, J 7.6 Hz, 7-H), 5.14
(4H, s, OCH2Ph), 6.77–6.79 (2H, m, ArH), 6.83–6.87
(3H, m, ArH), 6.88 (1H, d, J 1.8 Hz, ArH), 7.42–7.43
(4H, m, ArH), 7.26–7.39 (6H, m, ArH). NMR ꢁC
(CDCl3): 47.1, 52.4, 55.99, 56.01, 56.8, 60.3, 70.4, 71.0,
71.1, 77.4, 82.7, 94.0, 109.5, 110.4, 113.6, 113.9, 118.1,
120.3, 127.2, 127.3, 127.7, 127.9, 128.46, 128.53, 132.4,
135.1, 136.9, 137.2, 147.6, 148.3, 149.7, 150.0. Anal.
Found: C, 71.59; H, 6.59. Calcd. for C36H40O8: C,
71.98; H, 6.38%.
Measurement of the radical scavenging activity. To
an appropriate amount of a sample in a methanol solu-
tion (4.9 ml) was added 100 ml of 5 mM DPPH in a
methanol solution. After the solution had stood at 25 ꢀC
for 0.5 h, the absorbance at 517 nm was measured. The
antiradical activity was evaluated from the decreased
value of the 51-nm absorption, which was calculated by
the equation, % scavenging effect = ½ðA0 ꢁ A1Þ=A0ꢂ ꢃ
100, where A0 is the absorption of the control (without a
sample) and A1 is the absorption of the mixture
containing a sample.
(7S,70S,8R,80R)-4,40-Dibenzyloxy-3,30-dimethoxy-7,90:
70,9-diepoxy lignane (17). A reaction solution of MOM
ether 16 (0.64 g, 1.07 mmol) in THF (60 ml) and 6 M aq.
HCl solution (60 ml) was stirred at room temperature for
4 h before additions of H2O and EtOAc. The organic
solution was separated, washed with sat. aq. NaHCO3
solution, and brine, and dried (Na2SO4). Concentration
followed by silica gel column chromatography (EtOAc/
hexane = 1/3) gave furofuran 17 (0.44 g, 0.82 mmol,
Measurement of the superoxide scavenging activity.
The superoxide scavenging activity was determined by
using a WST superoxide dismutase assay kit (Dojindo
Molecular Technology, Kumamoto, Japan). Briefly,
superoxide radicals were generated by the xanthine/
xanthine oxidase system, and reduced WST-8 to water-
soluble formazane which exhibited an absorption max-
imum at 450 nm. The decreased absorption of the
reaction mixture indicated increased superoxide scav-
enging activity. The reaction mixture was incubated at
37 ꢀC for 20 min, and the absorption was read at 450 nm
by a Bio-Rad 680 microplate reader. The capability for
scavenging the superoxide radicals was calculated by
using the equation % scavenging effect = ½ðA0 ꢁ A1Þ=
77%) as a colorless crystals, mp 110–111 ꢀC (EtOH),
20
mp 108–110 ꢀC in the literature,9) ½ꢀꢂ
¼ þ15 (c 1.1,
D
CHCl3). The NMR data agreed with those in the lit-
erature.9) EIMS m=z: 538 (Mþ, 98), 91 (100). HREIMS
m=z: 538.2352 (calcd. for C34H34O6, 538.2356).
(7S,70S,8R,80R)-3,30-Dimethoxy-7,90:70,9-diepoxyli-
gnane-4,40-diol (pinoresinol) (5). Hydrogenolysis of 17
gave (+)-pinoresinol, mp 115–117 ꢀC, mp 118–120 ꢀC
20
in the literature,16) ½ꢀꢂ
¼ þ62 (c 0.2, CHCl3), ½ꢀꢂ ¼
D
D
þ64 (c 1.0, CHCl3) in the literature,16) The NMR data

2290
S. YAMAUCHI et al.
C.-T., and Lee, C. Y., American Chemical Society,
Washington DC, pp. 135–149 (1992).
A0ꢂ ꢃ 100, where A0 is the absorption of the control
(without a sample) and A1 is the absorption of the
mixture containing a sample.
8) Ward, R. S., and Hughes, D. D., Oxidative cyclization of
3,4-dibenzyltetrahydrofurans using ruthenium tetra(tri-
fluoroacetate). Tetrahedron, 57, 2057–2064 (2001).
9) Roy, S. C., Rana, K. K., and Guin, C., Short and stereo-
selective total synthesis of furano lignans (ꢄ)-dihydro-
sesamin, (ꢄ)-lariciresinol dimethyl ether, (ꢄ)-acumina-
tin methyl ether, (ꢄ)-sanshodiol methyl ether, (ꢄ)-
lariciresinol, (ꢄ)-acuminatin, and (ꢄ)-lariciresinol mon-
omethyl ether and furofuran lignans (ꢄ)-sesamin, (ꢄ)-
eudesmin, (ꢄ)-piperitol methyl ether, (ꢄ)-pinoresinol,
(ꢄ)-piperitol, and (ꢄ)-pinoresinol monomethyl ether by
radical cyclization of epoxides using a transition-metal
radical source. J. Org. Chem., 67, 3242–3248 (2002).
10) Yamauchi, S., Okazaki, M., Akiyama, K., Sugahara, T.,
Kishida, T., and Kashiwagi, T., First enantioselective
synthesis of (ꢁ)- and (+)-virgatusin, tetra-substituted
tetrahydrofuran lignan. Org. Biomol. Chem., 3, 1670–
1675 (2005).
Acknowledgments
This project was performed by grant-aid from 06
feasibility test related to corporation for innovative
technology and advanced research in evolutional area by
Toyo Industrial Creative Center. We measured the 400
MHz NMR data at INCS of Ehime University. Our
thanks to the president of Ehime University for sup-
porting this project. We are also grateful to Marutomo
Co. for financial support.
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