Isolation, synthesis, and biological activity of biphenyl and m-terphenyl-type compounds from Dictyostelium cellular slime molds

Article abstract of DOI:10.1016/j.tet.2012.08.041

From the fruiting bodies of Dictyostelium polycephalum, we obtained four aromatic compounds: dictyobiphenyl A (1) and B (2) and dictyoterphenyl A (3) and B (4). The synthesis of 1-4 was performed to confirm the structures and obtain sufficient material fo

Full text of DOI:10.1016/j.tet.2012.08.041

Tetrahedron 68 (2012) 8884e8889
Contents lists available at SciVerse ScienceDirect
Tetrahedron
journal homepage: www.elsevier.com/locate/tet
Isolation, synthesis, and biological activity of biphenyl and m-terphenyl-type
compounds from Dictyostelium cellular slime molds
,
a
a
b
a
Haruhisa Kikuchi a , Yusuke Matsuo , Yasuhiro Katou , Yuzuru Kubohara , Yoshiteru Oshima
*
a Graduate School of Pharmaceutical Sciences, Tohoku University, Aoba-yama, Aoba-ku, Sendai 980-8578, Japan
b Institute for Molecular and Cellular Regulation, Gunma University, Maebashi 371-8512, Japan
a r t i c l e i n f o
a b s t r a c t
Article history:
Received 29 June 2012
Received in revised form 11 August 2012
Accepted 13 August 2012
Available online 23 August 2012
From the fruiting bodies of Dictyostelium polycephalum, we obtained four aromatic compounds: dic-
tyobiphenyl A (1) and B (2) and dictyoterphenyl A (3) and B (4). The synthesis of 1e4 was performed to
confirm the structures and obtain sufficient material for biological evaluation. Compound 3 was the first
example of nitrogen-containing natural m-terphenyls, and the isolation of novel classes of compounds
such as 3 shows that cellular slime molds are promising sources in natural product chemistry. Moreover,
dictyoterphenyl A (3) showed cancer cell-selective antiproliferative activity (IC50 2.3e8.6 mM).
Keywords:
Cellular slime molds
m-Terphenyl
Ó 2012 Elsevier Ltd. All rights reserved.
Dictyostelium
Antiproliferative activity
Structural elucidation
1. Introduction
a resource for novel drug development, and we have studied the di-
versity of secondary metabolites of cellular slime molds.9 We have
The cellular slime mold Dictyostelium discoideum is thought to be
an excellent model organism for the study of cell and developmental
biology because of its simple pattern of development.1 Vegetative
cells of D. discoideum grow as single amoebae by eating bacteria.
When starved, they initiate a developmental program of morpho-
genesis and gather to form a slug-shaped multicellular aggregate.
This aggregate then differentiates into two distinct cell types, pre-
spore and prestalk cells, which are precursors of spores and stalk
cells, respectively. At the end of its development, the aggregate forms
a fruiting body consisting of spores and a multicellular stalk.
recently isolated a
-pyronoids,9a,f amino sugar derivatives,9c,d and aro-
matics9b,e,g with unique structures, and these compounds have dem-
onstrated several biological activities such as control of cellular slime
mold development. This paper reports the isolation, structure eluci-
dation, and synthesis of new biphenyl and m-terphenyl-type com-
pounds, dictyobiphenyl A (1), dictyobiphenyl B (2), dictyoterphenyl A
(3), and dictyoterphenyl B (4) from the cellular slime mold, Dictyoste-
lium polycephalum (Fig. 1). The selective antiproliferative activities of
these compounds in several cancer cell lines are also described.
Several small molecules including DIFs (differentiation-inducing
factors)2 and discadenine3 have been reported as development-
regulating substances of cellular slime molds. DIFs and their de-
rivatives also exhibit many biological effects in mammalian cells such
as suppression of cell growth,4e6 induction/promotion of cell differ-
entiation,4a,b promotion of glucose consumption,5 and regulation of IL-
2. Results and discussion
2.1. Isolation and structural elucidation
Fruiting bodies (wet weight 571 g) of D. polycephalum were
cultured on agar plates and extracted three times with methanol at
room temperature to yield an extract (19 g), which was partitioned
with ethyl acetate and water. The ethyl acetate solubles (4.0 g) were
separated by repeated column chromatography over silica gel and
ODS to yield 1 (3.8 mg), 2 (1.4 mg), 3 (2.0 mg), and 4 (0.5 mg).
HREIMS (m/z 243.0872 [M]þ) indicated that the molecular for-
mula of 1 was C14H13NO3 (Table 1). The 1H NMR spectrum indicated
2
production.6
A chlorine-substituted dibenzofuran derivative
AB0022A7 and a resorcinol derivative MPBD8 were also isolated from
cellular slime molds. However, except for these compounds, there have
been no reports on the secondary metabolites of cellular slime molds.
Thus, we have focused on the utility of cellular slime molds as
the presence of ABX spin system (
d
7.89 (1H, dd, J¼8.6, 2.2 Hz, H-4),
7.86 (1H, d, J¼2.2 Hz, H-2), and 7.11 (1H, d, J¼8.6 Hz, H-5)) and AB
* Corresponding author. Tel.: þ81 22 217 6824; fax: þ81 22 217 6821; e-mail
address: hal@mail.pharm.tohoku.ac.jp (H. Kikuchi).
spin system (
d
7.38 (H-20) and 6.86 (H-30) (each 2H, d, J¼8.7 Hz)) to
0040-4020/$ e see front matter Ó 2012 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.tet.2012.08.041

H. Kikuchi et al. / Tetrahedron 68 (2012) 8884e8889
8885
Fig. 2. Structural elucidation of dictyobiphenyl A (1).
1H NMR spectrum of 3 revealed the presence of additional ABX
spin system (
d
7.43 (1H, d, J¼2.2 Hz, H-200), 7.32 (1H, dd, J¼8.3,
2.2 Hz, H-600), and 6.97 (1H, d, J¼8.3 Hz, H-500)) along with the same
two spin systems as observed in 1 (Table 2). This fact implied the
existence of an additional 1,2,4-trisubstituted benzene ring (ring C).
The HREIMS of 3 (m/z 335.1148 [Mþ]) indicated a molecular formula
of C20H17NO4, which differed from that of 1 by C6H4O, suggesting
that the ring C contained a phenolic hydroxy group. The upfield
shift of the H-500 signal revealed that a hydroxy group was attached
at C-400. The HMBC correlation peaks for H-2eC-100, H-200eC-1, and
H-600eC-1 indicated that the benzene rings A and C were connected
through the C-1eC-100 bond. The rings B and C were connected
through the remaining C-300eC-10 bond. This connection was sup-
ported by the HMBC correlation between H-20 and C-300. Therefore,
the structure of 3 was elucidated (Fig. 3).
Fig. 1. Structures of dictyobiphenyl A (1) and B (2); and dictyoterphenyl A (3) and B (4).
Table 1
NMR spectral data of dictyobiphenyl A (1) and B (2)
Dictyobiphenyl A (1)a
Dictyobiphenyl B (2)b
13C
1H
13C
1H
1
2
3
4
5
6
7
8
10
130.9
129.8
Table 2
130.6 7.86 (1H, d, J¼2.2 Hz)
133.6 7.89 (1H, d, J¼2.1 Hz)
NMR spectral data of dictyoterphenyl A (3) and B (4)
127.5
123.2
128.8 7.89 (1H, dd, J¼8.6, 2.2 Hz) 131.0 7.79 (1H, dd, J¼8.4, 2.1 Hz)
Dictyoterphenyl A (3)a
Dictyoterphenyl B (4)b
111.7 7.11 (1H, d, J¼8.6 Hz)
116.7 6.89 (1H, d, J¼8.4 Hz)
159.9
169.1
160.0
170.3
13C
1H
13C
1H
1
2
3
4
5
6
7
8
10
130.9
129.8
56.0 3.85 (3H, s)
130.0
130.6 7.92 (1H, d, J¼2.3 Hz)
133.6 7.95 (1H, d, J¼2.0 Hz)
130.4
127.5
123.4
20, 60 131.5 7.38 (2H, d, J¼8.7 Hz)
131.4 7.39 (2H, d, J¼8.7 Hz)
115.9 6.82 (2H, d, J¼8.7 Hz)
157.8
129.0 7.90 (1H, dd, J¼8.6, 2.3 Hz) 131.0 7.80 (1H, dd, J¼8.3, 2.0 Hz)
30, 50 115.6 6.86 (2H, d, J¼8.7 Hz)
111.8 7.13 (1H, d, J¼8.6 Hz)
116.7c 6.92 (1H, d, J¼8.3 Hz)
40
157.6
160.0
168.9
160.0
170.4
a
600 MHz for 1H and 150 MHz for 13C in acetone-d6/methanol-d4 (9:1). Com-
56.1 3.87 (3H, s)
130.8
pound 1 was not possible to dissolve in only acetone-d6 or methanol-d4. Acetone-d6/
methanol-d4 (9:1) can sufficiently dissolve compound 1.
131.1
20, 60 131.3 7.48 (2H, d, J¼8.8 Hz)
131.5 7.45 (2H, d, J¼8.7 Hz)
b
600 MHz for 1H and 150 MHz for 13C in methanol-d4.
30, 50 115.7 6.86 (2H, d, J¼8.6 Hz)
115.9 6.83 (2H, d, J¼8.7 Hz)
40
157.3
157.4
100
200
300
400
500
600
130.5
130.8
imply a 1,2,4-trisubstituted benzene ring (ring A) and a 1,4-
disubstituted benzene ring (ring B) in 1, respectively (Fig. 2). The
13C NMR spectrum displayed 12 signals, and the DEPT spectra in-
dicated the existence of 6 quaternary carbons, 5 methine carbons,
and a methoxyl carbon. The difference between the number of 13C
NMR signals and the number of carbon atoms in the molecular
formula of 1 is two, supporting the presence of a symmetrical 1,4-
disubstituted benzene ring. The HMBC correlation peaks for H-
2eC-7, H-4eC-7, and H3-8eC-6 demonstrated that the carbonyl
group and the methoxy group were connected to C-1 and C-4, re-
spectively. The HMBC correlations for H-2eC-10 and H-20eC-1
suggested that the benzene rings A and B were connected through
the C-1eC-10 bond. The absorption at 1654 cmꢀ1 in the IR spectrum
of 1 revealed the existence of an aromatic amide, and an amido-
carbonyl group attached at C-3. Finally, the remaining hydroxy
group attached to C-40, elucidating the structure of 1.
132.3 7.43 (1H, d, J¼2.2 Hz)
132.4 7.45 (1H, d, J¼2.2 Hz)
128.8
154.2
129.6
154.5
116.3 6.97 (1H, d, J¼8.3 Hz)
116.6c 6.90 (1H, d, J¼8.4 Hz)
129.8 7.32 (1H, dd, J¼8.3, 2.2 Hz) 129.7 7.33 (1H, dd, J¼8.4, 2.2 Hz)
a
600 MHz for 1H and 150 MHz for 13C in acetone-d6/methanol-d4 (9:1). Com-
pound 3 was not possible to dissolve in only acetone-d6 or methanol-d4. Acetone-d6/
methanol-d4 (9:1) can sufficiently dissolve compound 3.
b
600 MHz for 1H and 150 MHz for 13C in methanol-d4.
These signals were indistinguishable.
c
HREIMS of 4 (m/z 322.0836 [M]þ) indicated a molecular formula
of C19H14O5. In comparison to 1H NMR, 13C NMR, and IR spectra of 3
and 4 (Table 2), compound 4 had a carboxy and hydroxy groups at
C-3 and C-6, respectively, in contrast to the amidocarbonyl and
methoxy groups observed in 3.
HREIMS of 2 (m/z 230.0583 [M]þ) indicated a molecular for-
mula, C13H10O4. The 1H and 13C NMR spectra of 2 were nearly
identical to those of 1 (Table 1), although compound 2 lacked the
signals of a methoxy group. Moreover, the absorption at 1683 cmꢀ1
in the IR spectrum of 2 was observed instead of the absorption at
1654 cmꢀ1 in 1. Therefore, compound 2 had a carboxy and hydroxy
groups at C-3 and C-6, respectively, in contrast to the amido-
carbonyl and methoxy groups detected in 1.
2.2. Synthesis
To confirm the structures and obtain a sufficient quantity of
material for performing several biological evaluations, we synthe-
sized compounds 1e4. Especially, the structures of terphenyl-type
compounds 3 and 4 had to be reconfirmed because their 13C NMR
signals between dC 127 and dC 133 were very crowded and difficult

8886
H. Kikuchi et al. / Tetrahedron 68 (2012) 8884e8889
Fig. 3. Structural elucidation of dictyoterphenyl A (3).
to assign. Preparation of rings B and C is shown in Scheme 1. A
SuzukieMiyaura coupling reaction was performed between 8 (the
ring C precursor) and pinacolatoboronate 11 (the ring B precursor)
to produce biphenyl 12 (ring CþB), which was then converted into
its triflate 13. The biphenyl skeleton of 12 was confirmed by its
HMBC correlations for H-2eC-10 and H-20eC-1.
Scheme 2. Synthesis of dictyobiphenyl A (1) and B (2).
Scheme 1. Synthesis of the precursors of rings B and C in 1e4.
Scheme 3. Synthesis of dictyoterphenyl A (3) and B (4).
In Scheme 2, dictyobiphenyl A (1) and B (2) were synthesized. A
2.3. Biological evaluation
palladium-catalyzed
boronation
of
methyl
3-bromo-4-
methoxybenzoate (15) produced compound 16, which corre-
sponded to ring A. Compounds 16 and 10 (ring B (Scheme 1)) were
linked by SuzukieMiyaura coupling reaction to give biphenyl 17
(ring AþB) in good yield. Although the amidation of 17 with
treatment of only ammonia was failed, the trimethylaluminum-
catalyzed amidation gave a good result, and the subsequent acidic
deprotection of MOM ether of 17 yielded dictyobiphenyl A (1). On
the other hand, treatment of 17 by BBr3 removed all MOM and
methyl groups to produce dictyobiphenyl B (2).
In Scheme 3, the coupling reaction of 13 (ring BþC) with 16 (ring
A) afforded terphenyl 18. Dictyoterphenyl A (3) and B (4) were
synthesized from 18 in a similar manner as the syntheses of 1 and 2,
respectively. All of the spectral data of synthetic 1e4 were identical
with those of natural 1e4, respectively, and thus their proposed
structures were confirmed.
The antiproliferative activities of 1e4 were evaluated on several
human cancer cell lines such as K562 leukemia, HeLa cervical
carcinoma, HCT116 colon carcinoma, and MCF-7 breast adenocar-
cinoma cells. In addition, the effects on mouse osteosarcoma LM8
cells (cancer cells) and mouse embryo fibroblast 3T3-L1 cells
(normal cells) were also evaluated. Compounds 1, 2, and 4 dem-
onstrated only weak activities and their IC50 values were more
than 20
proliferation of all cancer cell lines in a concentration dependent
manner, and its IC50 values were 8.6 M (HeLa), 2.3 M (K562),
6.9 M (HCT116), 5.4 M (MCF-7), and 4.9 M (LM8), respectively
(Fig. 4). On the other hand, the proliferation of 3T3-L1 cells did not
inhibit by up to 20 M of 3. These results indicated that dictyo-
mM for all cell lines. Dictyoterphenyl A (3) inhibited the
m
m
m
m
m
m
terphenyl A (3) showed cancer cell-selective antiproliferative
activity.

H. Kikuchi et al. / Tetrahedron 68 (2012) 8884e8889
8887
1.5% agar. When fruiting bodies had formed after 4 days, they were
harvested for extraction.
3.3. Isolation of dictyobiphenyl A (1) and B (2); and dictyo-
terphenyl A (3) and B (4)
The cultured fruiting bodies (dry weight 92.70 g) of D. poly-
cephalum were extracted three times with MeOH at room
temperature to give an extract (19.4 g), which was then parti-
tioned with EtOAc and H2O to yield EtOAc solubles (3.99 g). The
EtOAc solubles were chromatographed over silica gel, and the
column was eluted with hexane/EtOAc and EtOAc/MeOH solu-
tions with increasing polarity to afford hexane/EtOAc (1:3) el-
uent (fraction A, 134 mg). Fraction A was further separated
by ODS column using H2O/MeOH solvent system to give H2O/
MeOH (9:1) elutant (fraction B, 11 mg), dictyobiphenyl A (1)
(3.8 mg), and dictyoterphenyl
was subjected to recycle preparative HPLC (column, JAIGEL-
GS310 (
A (3) (2.0 mg). Fraction B
f
20 mmꢂ500 mm, Japan Analytical Industry Co., Ltd.);
solvent, MeOH) to give dictyobiphenyl B (2) (1.4 mg) and dic-
tyoterphenyl B (4) (0.5 mg).
3.3.1. Data for 1. Colorless amorphous solid; mp 248 ꢁC (dec); UV
(MeOH) lmax, nm (log )
) 205 (4.38), 248 (4.28); IR (KBr) nmax (cmꢀ1
3
3205, 2926, 1654, 1604; 1H NMR and 13C NMR data are shown in
Table 1; EIMS m/z 243 [M]þ (100%), 227 (53%); HREIMS m/z
243.0872 [M]þ (243.0895 calcd for C14H13NO3).
Fig. 4. Antiproliferative activity of dictyoterphenyl A (3) on HeLa and K562 cells (A),
HCT116 and MCF-7 cells (B), and LM8 cells (C). Cells were incubated in vitro with the
indicated concentrations of compound for several days, and the relative cell number
was assessed. The mean values and SD (bars) of the triplicate are presented.
3.3.2. Data for 2. Colorless amorphous solid; mp 208e210 ꢁC; UV
(MeOH) lmax, nm (log )
) 205 (4.42), 248 (4.39); IR (KBr) nmax (cmꢀ1
3
3277, 1683, 1605; 1H NMR and 13C NMR data are shown in Table 1;
EIMS m/z 230 [M]þ (100%), 213 (11%); HREIMS m/z 230.0583 [M]þ
(230.0579 calcd for C13H10O4).
Although many o-terphenyl-type compounds have ever been
reported in kingdom of fungi,10 very few m-terphenyl derivatives
such as dictyoterphenyl A (3) and B (4) occur naturally,10,11 in-
cluding dictyomedins previously reported by our group.9b In
particular, the isolation of dictyoterphenyl A (3) described in this
3.3.3. Data for 3. Colorless amorphous solid; mp 89e90 ꢁC; UV
(MeOH) lmax, nm (log ) 205 (4.61), 248 (4.57), 293 (sh, 4.00); IR
3
(KBr) nmax (cmꢀ1) 3354, 2926, 1653, 1604; 1H NMR and 13C NMR
data are shown in Table 2; EIMS m/z 335 [M]þ (100%), 317 (16%), 277
(26%), 243 (15%); HREIMS m/z 335.1148 [M]þ (335.1158 calcd for
C20H17NO4).
report represents the first-ever example for
a nitrogen-
containing natural m-terphenyl. Therefore, the isolation of
novel classes of compounds, such as 3 and 4, demonstrates that
cellular slime molds are promising sources in natural product
chemistry.
3.3.4. Data for 4. Colorless amorphous solid; mp 212e214 ꢁC; UV
(MeOH) lmax, nm (log ) 204 (4.51), 248 (4.47), 293 (sh, 3.87); IR
3
(KBr) nmax (cmꢀ1) 3293, 1685, 1605; 1H NMR and 13C NMR data are
shown in Table 2; EIMS m/z 322[M]þ (100%), 278 (8%); HREIMS m/z
322.0836 [M]þ (322.0841 calcd for C19H14O5).
3. Experimental section
3.1. General methods
3.4. 3-Bromo-1-(tert-butyl(dimethylsilyloxy))-4-(methox-
ymethoxy)benzene (8)
Analytical TLC was performed on silica gel 60 F254 (Merck).
Column chromatography was carried out on silica gel 60 (70e230
mesh, Merck). NMR spectra were recorded on JEOL ECA-600 and
AL-400. Chemical shifts for 1H and 13C NMR are given in parts per
To a solution of 712 (2930 mg, 9.66 mmol) in dichloromethane
(30 mL) were added chloromethyl methyl ether (0.95 mL,
12.5 mmol) and DIPEA (2.52 mL, 14.5 mmol) at 0 ꢁC. After being
stirred for 12 h at room temperature, the reaction mixture was
poured into saturated ammonium chloride solution, and extracted
with EtOAc three times. The combined organic layer was washed
with water and brine, dried over sodium sulfate, and evaporated.
The residue was chromatographed over silica gel eluted by hexane/
EtOAc (19:1) to give 8 (3100 mg, 8.93 mmol, 92%). Data for 8: col-
million (d) relative to tetramethylsilane (dH 0.00) and residual sol-
vent signals (dC 77.0, 29.8, and 49.0 for CDCl3, acetone-d6, and
CD3OD, respectively) as internal standards. Mass spectra were
measured on JEOL JMS-700 and JMS-DX303. IR spectra were mea-
sured on JASCO FT/IR-4200. UV spectra were measured on JASCO
V-550.
3.2. Organism and culture conditions
orless oil; 1H NMR (400 MHz, CDCl3)
(1H, d, J¼8.9 Hz), 6.71 (1H, dd, J¼8.9, 2.9 Hz), 5.15 (2H, s), 3.52 (3H,
s), 0.97 (9H, s), 0.18 (6H, s); 13C NMR (100 MHz, CDCl3)
151.0,148.5,
d
7.04 (1H, d, J¼2.9 Hz), 7.00
The cellular slime molds, D. polycephalum were kindly supplied
by Dr. Hagiwara, National Science Museum, Tokyo, Japan. Spores
were cultured at 22 ꢁC with Escherichia coli Br on A-medium con-
sisting of 0.5% glucose, 0.5% polypeptone, 0.05% yeast extract,
0.225% KH2PO4, 0.137% Na2HPO4$12H2O, 0.05% MgSO4$7H2O, and
d
124.7, 119.5, 117.5, 113.1, 96.0, 56.3, 25.6 (3C), 18.1, ꢀ4.5 (2C); LREIMS
m/z 348 [Mþ2]þ (100%), 346 [M]þ (97%), 291 (57%), 289 (55%), 267
(43%), 260 (17%), 258 (17%), 247 (22%), 245 (21%), 165 (20%);
HREIMS m/z 346.0590 [M]þ (346.0600 calcd for C14H23O3Br79Si).

8888
H. Kikuchi et al. / Tetrahedron 68 (2012) 8884e8889
3.5. 1-(Methoxymethoxy)-4-(4,4,5,5-tetramethyl-1,3,2-
dioxaborolan-2-yl)benzene (11)
was added to bis(pinacolato)diboron (228 mg, 0.898 mmol), [1,10-
bis(diphenylphosphino)ferrocene]palladium(II) dichloride dichlo
romethane adduct (66.7 mg, 0.082 mmol), and potassium acetate
(240 mg, 2.448 mmol) at room temperature. After being refluxed
for 18 h, the reaction mixture was evaporated. The residue was
chromatographed over silica gel eluted by hexane/EtOAc (9:1) to
give 16 (153 mg, 0.523 mmol, 64%). Data for 16: colorless oil; 1H
Under argon atmosphere, a solution of 1-bromo-4-(methox-
ymethoxy)benzene (10)13 (410 mg, 1.889 mmol) in dioxane (6 mL)
was added to bis(pinacolato)diboron (500 mg, 1.969 mmol), [1,10-
bis(diphenylphosphino)ferrocene]palladium(II) dichloride dichloro
methane adduct (146 mg, 0.179 mmol), and potassium acetate
(527 mg, 5.370 mmol) at room temperature. After being refluxed for
12 h, the reaction mixture was evaporated. The residue was chro-
matographed over silica gel eluted by hexane/EtOAc (19:1) to give 11
(442 mg, 1.672 mmol, 88%). Data for 11: colorless oil; 1H NMR
NMR (400 MHz, CDCl3)
d
8.34 (1H, d, J¼2.4 Hz), 8.09 (1H, dd,
J¼8.7, 2.4 Hz), 6.87 (1H, d, J¼8.7 Hz), 3.88 (6H, s), 1.36 (12H, s); 13C
NMR (100 MHz, CDCl3)
d 167.6, 166.8, 138.5, 134.5, 122.1, 109.8,
83.7 (2C), 55.9, 51.7, 24.8 (4C) (because a boron atom was bonded
to C-3, the C-3 signal was highly broadened and not observed);
LREIMS m/z 292 [M]þ (100%), 277 (19%), 261 (14%), 249 (31%), 235
(19%), 192 (37%); HREIMS m/z 292.1469 [M]þ (292.1482 calcd for
C15H21O5B).
(400 MHz, CDCl3)
d
7.75 (2H, d, J¼8.8 Hz), 7.02 (2H, d, J¼8.8 Hz), 5.20
(2H, s), 3.47 (3H, s), 1.33 (12H, s); 13C NMR (100 MHz, CDCl3)
d
159.7,
136.4 (2C),115.4 (2C), 94.0, 83.6 (2C), 56.0, 24.8 (4C) (because a boron
atom was bonded to C-4, the C-4 signal was highly broadened and
not observed); LREIMS m/z 264 [M]þ (100%), 234 (35%), 219 (11%);
HREIMS m/z 264.1527 [M]þ (264.1533 calcd for C14H21O4B).
3.9. Methyl 6-methoxy-40-(methoxymethoxy)biphenyl-3-
carboxylate (17)
3.6. 40,6-Bis(methoxymethoxy)biphenyl-3-ol (12)
Under argon atmosphere, a solution of 16 (71.2 mg, 0.291 mmol)
and 10 (70.8 mg, 0.268 mmol) in dioxane (2 mL) was added to [1,10-
bis(diphenylphosphino)ferrocene]palladium(II) dichloride dichloro
methane adduct (22.1 mg, 0.027 mmol), and tripotassium phos-
phate (175 mg, 0.824 mmol) at room temperature. After being
refluxed for 24 h, the reaction mixture was evaporated. The residue
was chromatographed over silica gel eluted by hexane-EtOAc (9:1)
to give 17 (75.4 mg, 0.249 mmol, 93%). Data for 17: colorless oil; 1H
Under argon atmosphere, a solution of 8 (562 mg, 1.618 mmol)
and 11 (329 mg, 1.24 mmol) in DMSO (6 mL) was added to [1,10-
bis(diphenylphosphino)ferrocene]palladium(II) dichloride dichloro
methane adduct (102 mg, 0.124 mmol), 1,10-bis(diphenylphosphino)
ferrocene (69.0 mg, 0.124 mmol), and potassium acetate (366 mg,
3.73 mmol) at room temperature. After being stirred for 20 h at
80 ꢁC, the reaction mixture was poured into water, and extracted
with EtOAc three times. The combined organic layer was washed
with water and brine, dried over sodium sulfate, and evaporated. The
residue was chromatographed over silica gel eluted by hexane/EtOAc
(4:1) to give 12 (144 mg, 0.496 mmol, 40%). Data for 12: colorless oil;
NMR (400 MHz, CDCl3)
7.09 (2H, d, J¼8.9 Hz), 6.97 (1H, d, J¼9.1 Hz), 5.21 (2H, s), 3.89 (3H,
s), 3.86 (3H, s), 3.50 (3H, s); 13C NMR (100 MHz, CDCl3)
166.8,
d
7.98e8.02 (2H, m), 7.46 (2H, d, J¼8.9 Hz),
d
160.1, 156.6, 132.2, 131.0, 130.6 (2C), 130.4, 130.0, 122.6, 115.8 (2C),
110.5, 94.4, 55.9, 55.7, 51.8; LREIMS m/z 302 [M]þ (100%), 272 (36%),
226 (11%); HREIMS m/z 302.1141 [M]þ (302.1154 calcd for
C17H18O5).
1H NMR (400 MHz, CDCl3)
d
7.42 (2H, d, J¼8.6 Hz), 7.06 (2H, d,
J¼8.6 Hz), 7.04 (1H, d, J¼8.7 Hz), 6.76 (1H, d, J¼3.1 Hz), 6.69 (1H, dd,
J¼8.7, 3.1 Hz), 5.67(1H, br s), 5.21 (2H, s), 4.97 (2H, s), 3.51 (3H, s), 3.36
(3H, s); 13C NMR (100 MHz, CDCl3)
d 156.3, 151.0, 148.0, 132.9, 131.8,
130.5 (2C), 118.2, 117.4, 115.8 (2C), 114.6, 96.1, 94.4, 56.01, 55.99;
LREIMS m/z 290 [M]þ (100%), 258 (39%), 213 (41%); HREIMS m/z
290.1135 [M]þ (290.1154 calcd for C16H18O5).
3.10. Synthesis of dictyobiphenyl A (1)
At 0 ꢁC, 2.0 M trimethylaluminium solution in toluene (387
0.774 mmol) was added to 0.5 M ammonia solution in dioxane
(774 L, 0.387 mmol). After 45 min, a solution of 17 (11.7 mg,
mL,
3.7. 40,6-Bis(methoxymethoxy)biphenyl-3-yl
trifluoromethanesulfonate (13)
m
0.039 mmol) in toluene (0.5 mL) was added to the reaction mix-
ture. After being stirred for 18 h at 60 ꢁC, the reaction mixture
was poured into 1 M hydrochloric acid, and extracted with EtOAc
three times. The combined organic layer was washed with satu-
rated sodium bicarbonate solution and brine, dried over sodium
sulfate, and evaporated. The residue was dissolved in 3% hydro-
gen chloride solution in MeOH (1.0 mL). After being stirred for 5 h
at room temperature, the reaction mixture was evaporated. The
residue was chromatographed over silica gel eluted by chloro-
To a solution of 12 (117 mg, 0.403 mmol) in dichloromethane
(3 mL) were added 2,6-lutidine (188
fluoromethanesulfonic anhydride (102
m
m
L, 1.61 mmol) and tri-
L, 0.606 mmol) at 0 ꢁC.
After being stirred for 4 h at room temperature, the reaction mix-
ture was poured into saturated sodium bicarbonate solution, and
extracted with EtOAc three times. The combined organic layer was
washed with water and brine, dried over sodium sulfate, and
evaporated. The residue was chromatographed over silica gel
eluted by hexane/EtOAc (49:1) to give 13 (146 mg, 0.346 mmol,
form/MeOH (19:1) to give dictyobiphenyl
0.028 mmol, 71% (two steps)).
A (1) (6.7 mg,
86%). Data for 13: colorless oil; 1H NMR (400 MHz, CDCl3)
d 7.43
(2H, d, J¼8.8 Hz), 7.25 (1H, d, J¼8.9 Hz), 7.20 (1H, d, J¼3.0 Hz), 7.15
(1H, dd, J¼8.9, 3.0 Hz), 7.13 (2H, d, J¼8.8 Hz), 5.22 (2H, s), 5.13 (2H,
3.11. Synthesis of dictyobiphenyl B (2)
s), 3.51 (3H, s), 3.42 (3H, s); 13C NMR (100 MHz, CDCl3)
d 157.0,
153.7, 143.9, 133.1, 130.5 (2C), 130.1, 123.3, 120.6, 118.8 (q, J¼321 Hz
(CF3)), 116.4, 116.0 (2C), 95.1, 94.4, 56.3, 56.1; LREIMS m/z 422 [M]þ
(100%), 346 (7%), 257 (13%); HREIMS m/z 422.0636 [M]þ (422.0647
calcd for C17H17O7F3S).
To a solution of 17 (18.0 mg, 0.060 mmol) in dichloromethane
(1 mL) was added boron tribromide (1.0 M solution in dichloro-
methane) (0.6 mL, 0.600 mmol) at ꢀ78 ꢁC. After being stirred for
30 min at ꢀ78 ꢁC and for additional 18 h at 0 ꢁC, the reaction
mixture was poured into 0.5 M hydrochloric acid, and extracted
with EtOAc three times. The combined organic layer was washed
with brine, dried over sodium sulfate, and evaporated. The resi-
due was chromatographed over silica gel eluted by chloroform/
MeOH (9:1) to give dictyobiphenyl B (2) (10.2 mg, 0.044 mmol,
76%).
3.8. Methyl 4-methoxy-3-(4,4,5,5-tetramethyl-1,3,
2-dioxaborolan-2-yl)benzoate (16)
Under argon atmosphere, a solution of methyl 3-bromo-4-
methoxybenzoate (15)14 (200 mg, 0.816 mmol) in dioxane (5 mL)

H. Kikuchi et al. / Tetrahedron 68 (2012) 8884e8889
8889
3.12. Methyl 6-methoxy-40,400-bis(methoxymethoxy)-m-ter-
phenyl-3-carboxylate (18)
absorbance at 450 nm was measured. A cell number was given as
a ratio of the absorbance (% of control).
In the same manner as the synthesis of 17, compound 18
(58.3 mg, 0.133 mmol, 43 %) was synthesized from 16 (114 mg,
0.390 mmol) and 13 (130 mg, 0.308 mmol). Data for 18: colorless
Acknowledgements
This work was supported in part by Grant-in-Aid for Scien-
tific Research (No. 23710247 and No. 24590110) from the Min-
istry of Education, Science, Sports and Culture of Japan, and the
SUNBOR GRANT from the Suntory Institute for Bioorganic
Research.
oil; 1H NMR (400 MHz, CDCl3)
d 7.99e8.03 (2H, m), 7.51 (2H, d,
J¼8.6 Hz), 7.48 (1H, d, J¼2.3 Hz), 7.46 (1H, dd, J¼8.6, 2.3 Hz), 7.26
(1H, d, J¼8.6 Hz), 7.10 (2H, d, J¼8.6 Hz), 6.99 (1H, d, J¼8.4 Hz), 5.22
(2H, s), 5.17 (2H, s), 3.89 (3H, s), 3.88 (3H, s), 3.51 (3H, s), 3.44 (3H,
s); 13C NMR (100 MHz, CDCl3)
d 166.9, 160.1, 156.3, 153.5, 132.2,
132.0, 131.9, 131.3, 130.9, 130.7 (2C), 130.5, 129.9, 129.3, 122.6, 115.7
(2C), 115.0, 110.5, 94.9, 94.4, 56.1, 56.0, 55.8, 51.9; LREIMS m/z 438
[M]þ (100%), 406 (38%), 361 (19%), 330 (44%), 299 (10%); HREIMS m/
z 438.1664 [M]þ (438.1679 calcd for C25H26O7).
Supplementary data
Supplementary data associated with this article can be found in
the online version, at http://dx.doi.org/10.1016/j.tet.2012.08.041.
These data include MOL files and InChiKeys of the most important
compounds described in this article.
3.13. Synthesis of dictyoterphenyl A (3)
References and notes
In the same manner as the synthesis of 1, dictyoterphenyl A (3)
(11.4 mg, 0.034 mmol, 67% (two steps)) was synthesized from 18
(22.2 mg, 0.051 mmol).
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In the same manner as the synthesis of 2, dictyoterphenyl B (4)
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fetal bovine serum (FBS), 25 mg/mL penicillin, and 50 mg/mL
streptomycin). HeLa and 3T3-L1 cells were in DMEM-HG (Dulbec-
co’s modified Eagle’s medium containing a high concentration
(4500 mg/L) of glucose supplemented with the antibiotics and 10%
FBS). LM8 cells were in MEMa (minimum essential medium alpha
modification supplemented with the antibiotics and 10% FBS). K562
(2ꢂ104 cells/well), LM8 (2.5ꢂ103 cells/well), 3T3-L1 (5ꢂ103 cells/
well), and HeLa (5ꢂ103 cells/well) were allowed to grow for 3
(K562, LM8, and 3T3-L1 cells) or 4 days (HeLa cells) in 12-well
plates; each well filled with 1 mL of the each medium containing
DMSO (0.2%) or sample compounds. The relative cell number was
assessed using Alamar blue (cell number indicator) as described
previously.4c
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presence or absence of drugs. After 3 days,10 mL of MTT reagent was
added to each well, and after 1 h incubation at 37 ꢁC (5% CO2),