First total synthesis of structurally unique flavonoids and their strong anti-inflammatory effect

Article abstract of DOI:10.1016/j.bmcl.2004.03.108

The first total synthesis of structurally unique flavonoids 1a and 1b is described. These compounds showed very strong anti-inflammatory effect against delayed hypersensitivity in a mouse model.

Full text of DOI:10.1016/j.bmcl.2004.03.108

Bioorganic & Medicinal Chemistry Letters 14 (2004) 3201–3203
First total synthesis of structurally unique flavonoids and their
strong anti-inflammatory effect
Takashi Nakatsuka,a,* Yoshiaki Tomimori,a Yoshiaki Fukudaa and Haruo Nukayab
aDaiichi Suntory Biomedical Research Co., Ltd, 1-1-1, Wakayamadai, Shimamoto-cho, Mishima-gun, Osaka 618-8513, Japan
bSchool of Pharmaceutical Science, University of Shizuoka, 52-1 Yada, Shizuoka 422-8526, Japan
Received 11 March 2004; accepted 31 March 2004
Abstract—The first total synthesis of structurally unique flavonoids 1a and 1b is described. These compounds showed very strong
anti-inflammatory effect against delayed hypersensitivity in a mouse model.
Ó 2004 Elsevier Ltd. All rights reserved.
Flavonoids, including apigenin, luteolin, diosmin, and
rutin show diverse biological properties1 such as anti-
inflammatory, anti-cancer, and anti-oxidant effects.
Recently, Ishikura et al. discovered a novel and struc-
turally unique flavonoid (1a) from oolong tea extract,
which showed very strong anti-inflammatory activity.2
Although structurally related flavone C-glycosides such
as vitexin and isovitexin are widespread in nature and a
synthetic method was also investigated,3 a fused tricyclic
flavone, such as 1, has not been found.
O
O
HO
R3O
Ar
O
O
OH
O
HO
HO
R2O
R2O
O
O
OH
OR2
1a : Ar=p-OH-C6H4
1b : Ar=C6H5
2
O
OR4
R3O
O
X
R2O
R2O
In our effort to exploit a new anti-inflammatory agent,
we are very interested in 1a and its derivatives, but the
content of 1a in oolong tea is very low. In addition, there
are no reports of synthetic methods for this type of
compound. Here, we wish to report the first total
synthesis of 1a and its derivative (1b) and their strong
anti-allergic activity against contact and delayed
hypersensitivity in a mouse model.
O
R2O
R2O
OR1
OH
OR2
OH
OR2
3
4
Scheme 1. Retrosynthetic analysis.
epoxidation6 using dimethyldioxirane, followed by
treatment with methanol, methyl 3,4,6-tri-O-benzyl-b-
The retrosynthetic analysis of these compounds is pre-
sented in Scheme 1. As one can observe, the key factors
of this synthesis are the selection of protective groups of
sugar and phenolic hydroxyl groups and the cyclization
stepof diol 3 to fused tricyclic compound 2. Previously
reported methodologies were applied to the synthesis of
C-glycoside 3 from 44 and the formation of flavone 1
from acetophenone derivative 2.5
D
-
glucoside was obtained. For acid stability and a depro-
tective condition, p-nitrobenzyl ether was selected as a
protective group7 for the 2-position of this glucose
derivative. After hydrolysis of methyl glucoside with dil
H2SO4, then conversion to the corresponding glycosyl
fluoride (7) using (diethylamino)sulfur trifluoride
(DAST) at )78 °C,8 C-glycosidation of 7 with 1-[2,4-
bis(benzyloxy)-6-hydroxyphenyl]ethanone (15)9 was
examined using several reported methods,4 and
BF3OEt2/MS4A in CH2Cl2 gave the best results. Inter-
estingly, the protective groups of both the glycosyl donor
and acceptor greatly affected the C-glycosidation results.
Among the hydroxyl protective groups of the 2-position
As shown in Scheme 2, we chose 3,4,6-tri-O-benzyl-D-
glucal (5) as a starting material. After stereoselective
* Corresponding author. Tel.: +81-75-962-8184; fax: +81-75-962-6448;
e-mail: takashi_nakatsuka@dsup.co.jp
0960-894X/$ - see front matter Ó 2004 Elsevier Ltd. All rights reserved.
doi:10.1016/j.bmcl.2004.03.108

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T. Nakatsuka et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3201–3203
Scheme 2. Reagents and conditions: (a) dimethyldioxirane, CH2Cl2, 0 °C, quant; (b) MeOH/CH2Cl2, rt, quant.; (c) p-NO2–C6H4CH2Br, 2,4,6-
collidine, AgOTf, 35 °C, 63%; (d) 2 M H2SO4, AcOH, reflux, 50%; (e) DAST, CH2Cl2, )78 °C, 78%; (f) 1-[2,4-bis(benzyloxy)-6-hydroxy-
phenyl]ethanone (15), BF3OEt2, MS4A, )20 °C, then rt, 56%; (g) MeI, NaH, DMF, rt, 86%; (h) indium powder, satd NH4Cl, MeOH/iPrOH, 85 °C,
73%; (i) 10% Pd–C, THF, rt, 92%; (j) 1,10-azobis(N,N-dimethylformamide), Bu3P, benzene, rt, 73%; (k) p-benzyloxybenzoyl chloride, DMAP,
CH2Cl2, rt, 88%; (l) LDA, THF, )30 °C, 50%; (m) TMSOTf, CH2Cl2, rt, 62%; (n) BCl3, CH2Cl2, rt, then NaHCO3, HPLC purification, 10%; (o)
PhCHO, 50% NaOH, dioxane, rt, 50%; (p) C6H5I(OAc)2, KOH/MeOH, rt, 52%; (q) BCl3, CH2Cl2, rt, evaporation, then aq 1 M HCl–MeOH; rt,
3 days, 44%.
of 4, such as p-nitrobenzyl ether, triethylsilyl ether, and
methoxymethyl ether, only p-nitrobenzyl ether gave the
desired product. Among the protective groups of 2,4,6-
trihydroxyacetophenone, such as benzyl ether, t-butyl-
diphenylsilyl ether, and t-butyldimethylsilyl ether, only
15 gave the desired product 8. After protection of the
phenolic hydroxyl group of 8 by MeI and NaH, followed
by deprotection of the p-nitrobenzyl groupby indium–
aqueous ammonium chloride,5b and deprotection of
phenolic benzylether by Pd–C/H2 in THF, 9 was ob-
tained. With cyclization of triol (9) by the Mitsunobu
condition,10 the desired compound 10 was obtained in
good yield. Using this key intermediate, the flavones 12
and 14 were synthesized in two different ways. The nat-
ural product precursor (12) was synthesized by acylation
of 20-OH by acyl chloride/DMAP, then rearrangement of
the acyl groupby a large excess of LDA, and finally,
cyclization by TMSOTf.3 On the other hand, 14 was
prepared by condensation with benzaldehyde, followed
by flavone formation with hypervalent iodine reagent.5b
Debenzylation of 14 using Pd–C/H2 in AcOEt did not
give the desired deprotected compound but gave a furan
ring-decomposed compound. To obtain the desired
product, fully protected 12 or 14 were treated with BCl3
in CH2Cl2 at room temperature for 5 min, then the
reaction was quenched by aqueous NaHCO3 or MeOH,
respectively. Because boronic ester along with the desired
product was obtained in the case of MeOH quenching,
the crude reaction mixture was treated with aqueous 1N–
HCl/MeOH at room temperature for 3 days to give the
desired product 1b. By HPLC purification, pure 1a and
1b11 were obtained.
Oral administration of chemically synthesized 1b
reduced the skin reaction, which was elicited by an
epicutaneous application of 2,4,6-trinitro-1-chlorobenz-
ene (TNCB) to the ears of sensitized mice in a dose-
dependent manner at 2 and 10 lg/kg, as shown in Figure
1.12 Compared with natural product (1a), which reduced
the increase of ear thickness by 30% at a dose of 10 lg/
kg, 1b showed stronger effect (59% reduction at the same
dose). And compared with betamethasone, which was
#
150
*
100
* *
* *
50
0
control
vehicle
2 mg/kg
0.4
2
10 µg/kg
Betamethasone
compound 1b
Valerate
Figure 1. Effect of 1b on contact hypersensitivity reaction in mice.
Values are means SEM (N ¼ 7). Contact hypersensitivity reaction
was induced by topical application of TNCB to the ears of sensitized
mice, and the skin reaction was determined at 24 h after the elicitation.
(#) P < 0:01 compared with unsensitized control group (Student’s t-
test); (*) P < 0:05; (**) P < 0:01 compared with vehicle group
(Dunnett’s test).

T. Nakatsuka et al. / Bioorg. Med. Chem. Lett. 14 (2004) 3201–3203
3203
6. Halcomb, R. L.; Danishefsky, S. J. J. Am. Chem. Soc.
1989, 111, 6661.
administered orally at a dose of 2 mg/kg, the efficacious
dose of 1b was much lower.
7. (a) Fukase, K.; Tanaka, H.; Torii, S.; Kusumoto, S.
Tetrahedron Lett. 1990, 31, 389; (b) Moody, C. J.; Pitts,
M. R. Synlett 1999, 1575.
8. (a) Rosenbrook, W., Jr.; Riley, D. A.; Lartey, P. A.
Tetrahedron Lett. 1985, 26, 3; (b) Posner, G. H.; Haines,
S. R. Tetrahedron Lett. 1985, 26, 5.
9. Kumazawa, T.; Minatogawa, T.; Matsuba, S.; Sato, S.;
Onodera, J. Carbohydr. Res. 2000, 329, 507.
10. (a) Mitsunobu, O.; Yamada, M. Bull. Chem. Soc. Jpn.
In conclusion, we have demonstrated the first total
synthesis of tricyclic flavonoid 1a and its application to
the synthesis of a new derivative 1b, which showed a
stronger anti-inflammatory effect than that of the nat-
ural flavonoid 1a.
Further studies on the mechanistic aspects of the anti-
inflammatory effects of these novel flavonoids are under
investigation.
^
1967, 40, 2380; (b) Tsunoda, T.; Yamamiya, Y.; Ito, S.
Tetrahedron Lett. 1993, 34, 1639.
11. 1H NMR (400 MHz, CD3OD): d 3.3–3.4 (m, 1H), 3.55–
3.65 (m, 2H), 3.82 (dd, 1H, J ¼ 3 Hz, 12 Hz), 3.99 (dd, 1H,
J ¼ 5 Hz, 9 Hz), 4.7–4.75 (m, 1H), 5.22 (d, 1H, J ¼ 3 Hz),
6.70 (s, 1H), 6.81 (s, 1H), 7.5–7.6 (m, 3H), 7.95–8.05 (m,
2H).
Acknowledgements
12. Asherson, G. L.; Ptak, W. Immunology 1968, 15, 405.
Female BALB/c mice (7–9 weeks old, N ¼ 7, Charles
River Japan, Inc., Kanagawa) were sensitized by an
epicutaneous application with 100 lL of 7% 2,4,6-trini-
tro-1-chlorobenzene (TNCB) (in a 4:1 mixture of acetone
and olive oil) to shaved abdomen of mice. Six days after
the sensitization, 20 lL of 1% TNCB in acetone/olive oil
(1:9) was applied to each side of right ears of the mice to
induce contact hypersensitivity response. As a control, the
TNCB solution was painted to ears similarly to unsensi-
tized mice. Test compounds were suspended in 0.5% of
hydroxy propyl cellulose (HPC) (Nippon Soda Co., Ltd,
Tokyo, Japan) and administered orally three times (30 min
prior to the elicitation, 6 and 21 h after the elicitation). Ear
thickness was measured at 0 and 24 h after the elicitation
utilizing a thickness gauge (Digimatic Indicator, Mitsu-
toyo, Tokyo, Japan), and contact hypersensitivity
response was determined by the difference. Betamethasone
valerate (Wako pure chemicals, Osaka, Japan) was
suspended in 0.5% HPC and administered orally as a
positive control. The statistical analysis was performed
with Dunnett’s multiple comparison test or Student’s t-test
using SuperANOVA (Abacus Concepts, Berkeley, CA) or
Statview (SAS Institute Inc., Cary, NC), respectively. The
P-value less than 0.05 was considered significant.
We are grateful to Professor Kuniro Tsuji (University of
Shizuoka) for helpful discussion. Special thanks to Mr.
H. Inoue, Dr. T. Tomoo, Dr. T. Muto, Mr. T. Tanaka,
Mr. H. Murafuji, and Ms. J. Futamura for their
involvement in the project.
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5. (a) Gripenberg, J. In The Chemistry of Flavonoid Com-
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ꢀ
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