Synthesis and inhibition of PGE2 production of 6,8-disubstituted chrysin derivatives

Article abstract of DOI:10.1016/j.ejmech.2005.04.013

A series of 6,8-disubstituted chrysin derivatives have been synthesized and evaluated for their PGE₂ inhibitory activities. 6,8-Disubstituted chrysin derivatives were obtained from naturally occurring chrysin by halogenation, oxidation, thiomet

Full text of DOI:10.1016/j.ejmech.2005.04.013

European Journal of Medicinal Chemistry 40 (2005) 943–948
www.elsevier.com/locate/ejmech
Short communication
Synthesis and inhibition of PGE2 production
of 6,8-disubstituted chrysin derivatives
Haeil Park *, Tran Thanh Dao, Hyun Pyo Kim
College of Pharmacy, Kangwon National University, Chuncheon 200-701, South Korea
Received 29 June 2004; received in revised form 8 April 2005; accepted 28 April 2005
Available online 15 June 2005
Abstract
A series of 6,8-disubstituted chrysin derivatives have been synthesized and evaluated for their PGE2 inhibitory activities. 6,8-Disubstituted
chrysin derivatives were obtained from naturally occurring chrysin by halogenation, oxidation, thiomethylation and C–C cross coupling
reaction. Among the compounds investigated, 6,8-dibromochrysin (2), 6,8-diiodochrysin (4), 6,8-dimethylthiochrysin (9) and 6,8-
dimethoxychrysin (11) showed as strong inhibitory activities of PGE2 production from LPS-induced RAW 264.7 cells as wogonin, a well
known natural flavone having strong and selective COX-2 inhibitory activity.
© 2005 Elsevier SAS. All rights reserved.
Keywords: Chrysin; PGE2 production; Anti-inflammatory; COX-2; Wogonin; Flavonoids
1. Introduction
taglandin production [8,9]. The results showed that methyla-
tion of 5,7-dihydroxy groups on the A ring as well as substi-
tutions on the B ring of chrysin did not affect to the bioactivity.
As part of our continuing research efforts directed at the SARs
of natural flavones for the anti-inflammatory activity, we were
interested in the effects of hydrophobic groups such as meth-
ylthio (–SCH3), halogen (I, Br), alkyl and aryl groups substi-
tuted at the 6- and 8-positions of chrysin. Based on the struc-
ture–activity relationships (SARs) of these compounds for
anti-inflammation, we visualized that the substitutions either
at 6- or 8-position on the A ring of chrysin may play impor-
tant roles in their bioactivities. These substituents are rarely
seen in the basic structure of natural flavones, but it would be
believed that they have important roles of bioactivities. In
this paper, the synthesis of 6,8-disubstituted chrysin deriva-
tives and evaluation of their inhibitory activities on PGE2 pro-
duction are reported. These compounds are more structurally
similar to wogonin and oroxylin A than chrysin. As observed
from the inhibitory activities of PGE2 production of wogo-
nin, baicalein and oroxylinA, substituention at 6- or 8-position
did increase the bioactivity compared to that of chrysin. These
results led us to design chrysin derivatives with substituents
at 6- and 8-position. All the compounds were easily obtained
from chrysin with excellent yields in a few steps.
Chrysin (5,7-dihydroxyflavone), a naturally wide distrib-
uted flavonoid, has been reported to possess diverse biologi-
cal activities such as anti-oxidant, anti-allergy, anti-
inflammatory and anti-cancer [1–4]. It has been proposed that
chrysin acts as an agonist of PPAR-c which results in down-
regulation of the key pro-inflammatory enzymes, inducible
nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2)
[5]. Furthermore, some natural flavone analogs such as wogo-
nin, baicalein and oroxylin A (Fig. 1) showed much stronger
inhibitory activities of PGE2 production than that of chrysin
[6,7]. These flavones have an extra 6- or 8-substituent group
at the A ring of the flavone structures compared to the struc-
ture of chrysin. Therefore, we assumed that structural modi-
fication at both 6- and 8-positions of the A ring of chrysin
could be tolerable to the bioactivity.
As an attempt to discover novel synthetic flavones with
potent anti-inflammatory activity, recently we have synthe-
sized flavone analogs modified at theA and B ring systems of
chrysin and evaluated their inhibitory activities against pros-
* Corresponding author. Tel.: +82 33 250 6920; fax: +82 33 255 7865.
E-mail address: haeilp@kangwon.ac.kr (H. Park).
0223-5234/$ - see front matter © 2005 Elsevier SAS. All rights reserved.
doi:10.1016/j.ejmech.2005.04.013

944
H. Park et al. / European Journal of Medicinal Chemistry 40 (2005) 943–948
Fig. 1. Structures of some natural flavone analogs.
2. Chemistry
carbonate gave 5,7-dimethoxy-6,8-dibromoflavone (3),
5-hydroxy-7-methoxy-6,8-diiodoflavone (5), and 6,8-diiodo-
5,7-dimethoxyflavone (6). Suzuki coupling reaction [12] of
6, benzeneboronic acid and catalytic amount of Pd(PPh3)4
afforded 5,7-dimethoxy-6,8-diphenylflavone (7). Deprotec-
tion reaction of 7 with BBr3 in anhydrous methylene chloride
gave 5,7-dihydroxy-6,8-diphenylflavone (8). Thiomethyla-
tion reaction [13] of chrysin (1) gave 6,8-dimethylthiochrysin
(9) and followed by methylation with dimethylsulfate yielded
The synthetic procedures and reaction conditions for all
investigated compounds are shown in Schemes 1 and 2.
Chrysin (1) was directly halogenated by process of bromina-
tion [10] to form 6,8-dibromochrysin (2), or iodination [11]
to form 6,8-diiodochrysin (4). Methylation reaction of the 6,8-
dihalogenated chrysin derivatives with either 1 or 2 M equiva-
lents of dimethylsulfate in anhydrous acetone and potassium
Scheme 1. i: Bromine, CHCl2, Me2S, 0 °C; ii: Me2SO4 (2 equiv.), K2CO3, acetone, reflux; iii: iodine, acetic acid, 0 °C; iv: Me2SO4 (1 equiv. For 5; 2 equiv. for
6), K2CO3, acetone, reflux; v: benzeneboronic acid, DMF, Pd (PPh3)4, 90 °C; vi: BBr3, chloroform, reflux; vii: DMDS, FeCl3, toluene, reflux; viii: Me2SO4
(2.5 equiv.), K2CO3, acetone, reflux.

H. Park et al. / European Journal of Medicinal Chemistry 40 (2005) 943–948
945
Scheme 2. i: Benzyl bromide, K2CO3, acetone, 60 °C; ii: K2S2O8, pyridine, KOH; iii: Me2SO4, K2CO3, acetone 60 °C; iv: c-HCl, acetic acid, 80 °C; v: c-HCl,
acetic acid, 80 °C; vi: Me2SO4, K2CO3, acetone 60 °C.
6,8-dithiomethyl-5,7-dimethoxyflavone (10). Reactions of
chrysin via benzylation, Elbs persulfate oxidation [14],
methylation and debenzylation yielded 5,6,7,8-tetra-
hydroxyflavone (11), 5,6,7,8-tetra-methoxyflavone (12) and
5,7-dihydroxy-6,8-dimethoxyflavone (13) as described in
Scheme 2. The synthesized compounds were purified by flash
column chromatography and analyzed the structures based
on 1H-NMR and 13C-NMR spectra.
5- and 7-positions (2, 4, 9 and 11) generally showed very
strong inhibitory activities of COX-2 catalyzed PGE2 produc-
tion as shown in Table 1. While all 6,8-disubstituted chrysin
derivatives with methoxy group(s) at 5- and/or 7-position(s)
(3, 5, 6, 7, 10 and 13) showed significantly decreased bioac-
tivities. The compound having hydroxy groups at 6- and
8-positions (12) showed much reduced bioactivity though it
has the free hydroxy groups at 5- and 7-positions. The com-
pounds having phenyl substituents at 6- and 8-positions (7
and 8) showed moderate activities regardless of the free
hydroxy groups at 5- and 7-positions. These results may imply
that the character of substituents at the 6- and 8-positions is
related with bioactivities of chrysin derivatives. Based on our
3. Pharmacology
The bioassays were performed according to the published
procedure [7]. RAW 264.7 cells obtained fromAmerican Type
Culture Collection were cultured with DMEM supplemented
with 10% FBS and 1% CO2 at 37 °C and activated with LPS
(Lipopolysaccharide, Escherichia coli O127:B8). Briefly,
cells were plated in 96-well plates (2 × 105 cells per well).
Each synthetic flavone was dissolved in dimethyl sulfoxide
(DMSO) and LPS (1 µg/ml) were added and incubated for
24 h. Cell viability was assessed with MTT assay based on
the experimental procedures described previously [15]. All
tested compounds showed no or less than 10% reduction of
MTT assay, indicating that they were not significantly cyto-
toxic to RAW 264.7 cells in the presence or absence of LPS.
PGE2 concentration in the medium was measured using EIA
kit for PGE2 according to the manufacturer’s recommenda-
tion. All experiments were carried out at least twice and they
gave similar results. The inhibitory activities of synthetic fla-
vones on COX-2 catalyzed PGE2 production from LPS-
induced RAW 264.7 cells were estimated and shown in
Table 1.
Table 1
Inhibition of COX-2 catalyzed PGE2 production from LPS-induced RAW
264.7 cells by synthetic chrysin derivatives
Flavonesa
Percent inhibition of
PGE2 productionb
11.12
IC50 (µM)
1 (Chrysin)
2
3
4
5
6
NTc
1.34
NT
99.93
Inactive
98.41
1.82
NT
Inactive
57.13
NT
7
43.73
NT
8
39.38
NT
9
10
11
12
98.53
0.96
NT
Inactive
99.20
2.85
NT
22.43
13
64.78
NT
Wogonin
99.27
1.08
a All compounds were treated at 10 µM. Treatment of LPS to raw cells
increased PGE2 production (10 µM) from the basal level of 0.5 µM.
b % Inhibition = 100 × [1 – (PGE2 of LPS with the flavones treated group
– PGE2 of the basal)/(PGE2 of LPS treated group – PGE2 of the basal)].
c NT: not tested.
4. Results and discussion
Among the synthesized 6,8-disubstituted chrysin deriva-
tives, the compounds which have free hydroxy groups at the

946
H. Park et al. / European Journal of Medicinal Chemistry 40 (2005) 943–948
present results, it may be concluded that the 5,7-dihydroxy
groups as well as the character of substituents at 6- and
8-positions on the A ring of chrysin derivatives play impor-
tant roles on the inhibition of PGE2 production. The active
compounds with inhibition values over 90% were tested for
MTT assay. The results indicated that all tested compounds
did not exhibit any cytotoxicity. Therefore, the inhibition of
PGE2 production by flavone derivatives were not associated
with their cytotoxicity.
In summary, we prepared chrysin derivatives modified at
the 6- and 8-positions of theA ring and evaluated their inhibi-
tory activities on COX-2 catalyzed PGE2 production from
LPS-induced RAW 264.7 cells. We found that 6,8-dihalogeno
(2 and 4), 6,8-dimethylthio (10) and 6,8-dimethoxy (11)
chrysin derivatives with 5,7-dihydroxy groups possessed
strong inhibitory activities compared to that of chrysin. The
present results indicated that the free hydroxyl groups at 5-
and 7-potitions and the character of substituents at the 6- and
8-positions of theA ring play very important roles to biologi-
cal activity as we premised. The strong inhibitory activities
of PGE2 production by 6,8-disubstituted chrysin derivatives
may be explained by the structural similarity of these com-
pounds, wogonin and oroxylin A. In conclusion, substitution
at both 6- and 8-posiotion is tolerable to bioactivity but size
and electronic character of the substituents seem to closely
related with bioactivity. Our present results are quite differ-
ent from the previous results by us that methylation of 5,7-
dihydroxy groups on the A ring of chrysin did not affect to
the bioactivity [8]. Further SARs study on the A ring is cur-
rently under investigation.
After cooling in ice-water bath, a solution of 1.3 ml of bro-
mine in 20 ml of CH2Cl2 was added slowly with magnetic
stirring over 10 min. Then, a solution of 5.08 g of chrysin in
50 ml of CH2Cl2 was added slowly over a few min., and stir-
ring was continued for another 2 h and 30 min. At the end of
reaction (monitoring by TLC with the solvent system of chlo-
roform and methanol 20:1), the solvent was removed by
evaporation in vacuum, followed by treatment with saturated
NaHCO3 solution to remove trace of HBr. The remain solid
was washed with water and crystallized from absolute metha-
1
nol to obtain the title compound. Yield 92%. H-NMR
(200 MHz, DMSO-d6): d 13.75 (s, 1H, OH), 11.77 (s, 1H,
OH), 8.12–8.16 (d, 2H, H2’, H6’), 7.60–7.64 (t, 3H, H3’,
H4’, H5’), 7.21 (s, 1H, H3). 13C-NMR (50 MHz, DMSO-d6):
d 181.5 (C-4), 163.4 (C-7), 157.9 (C-2), 157.1 (C-10), 152.4
(C-5), 132.5 (C-1’), 130.3 (C-3’and C-5’), 129.3 (C-4’), 126.5
(C-2’ and C-6’), 105.1 (C-9), 104.8 (C-3), 94.7 (C-6), 88.6
(C-8). EIMS: m/z 413 (M + 1). Anal. C15H8Br2O4 (C, H, N).
5.1.2. 6,8-Dibromo-5,7-dimethoxyflavone (3)
A mixture of 6,8-dibromo-5,7-dihydroxyflavone (1 equiv.)
and K2CO3 and dimethyl disulfate (2.2 equiv.) in acetone was
refluxed for 6 h, monitoring by TLC. The mixture was fil-
tered and filtration was concentrated. Solidified product was
filtered, washed with water, and recrystallized from metha-
nol to form the titled flavone. 1H-NMR (200 MHz, CDCl3): d
8.18–8.24 (d, 2H, H2’, H6’); 7.68–7.75 (m, 3H, H3’, H4’,
H5’); 7.17 (s, 1H, H3); 3.94-4.03 (s, 6H, 2xOCH3). 13C-
NMR (50 MHz, DMSO-d6): d 175.8 (C-7), 166.5 (C-4), 161.7
(C-5), 160.8 (C-2), 153.5 (C-9), 132.2 (C-1’), 130.4 (C-3’
and C-5’), 129.4 (C-4’), 126.3 (C-2’and C-6’), 115.6 (C-10),
108.1 (C-3), 99.0 (C-8), 77.6 (C-6), 61.66 and 61.00 (2 ×
OCH3). EIMS: m/z 440 (M+). Anal. C17H12Br2O4 (C, H, N).
5. Experimental section
5.1. Chemistry
5.1.3. 5,7-Dihydroxy-6,8-diiodoflavone (4)
All chemicals were purchased from commercial suppli-
ers, and used without further purification. All solvents used
for reaction were freshly distilled from proper dehydrating
A mixture of iodine (20 mmol) in 20 ml of CH2Cl2 and
5,7-dihydroxyflavone (20 mmol) in 20 ml of glacial acetic
acid was stirred at room temperature for 30 min. during the
slow addition of 2 g of 65% HNO3 acid in 10 ml of acetic
acid. The iodinated product was precipitated during the addi-
tion. The suspension was stirred for 2 h and filtered. The solid
was washed with 10% Na2S2O4 solution then cooled metha-
nol and water. The product was crystallized as pale yellow
crystals.Yield 75%. 1H-NMR (200 MHz, DMSO-d6): d 13.89
(s, 1H, OH), 8.08–8.12 (m, 2H, 6.8 Hz, H2’, H6’), 7.48–7.53
(m, 3H, H3’, H4’), 7.08 (s, 1H, H3). 13C-NMR (50 MHz,
DMSO-d6): d 181.5 (C-4), 178.5 (C-7), 163.8 (C-2), 162.0
(C-5), 161.1 (C-10), 132.6 (C-1’), 130.4 (C-3’and C5’), 129.4
(C-4’), 126.8 (C-2’and C-6’), 105.1 (C-9), 104.7 (C-3), 63.4
(C-6 and C-8). EIMS: m/z 506 (M+). Anal. C15H8I2O4 (C, H,
N).
1
agents under nitrogen gas. H-NMR spectra were recorded
on a Varian Gemini 2000 instrument (200 MHz) spectrom-
eter. 13C-NMR spectra were recorded on a Varian Gemini
2000 instrument (50 MHz) spectrometer. Chemicals shifts are
reported in parts per million (ppm) downfield relative to tet-
ramethylsilane as a internal standard on the d scale. Data are
reported as follows: chemical shift, multiplicity (s = single,
d = double, t = triplet, q = quartet, m = multiplet, br = broad),
coupling constant (Hz), integration. Chromatographic purifi-
cation was carried out by flash chromatography using Kiesel-
gel 60 (230–240 mesh, Merck). Mass spectra were recorded
on GCMS (Autospec. M363 series) auto sampler/direct injec-
tion (EI). Elemental analyses were recorded on Series II CHN
s/o Analyzer PE 2400 AD:6 and the data for C, H, N were
with in 0.4% of the theoretical values.
5.1.4. 5-Hydroxy-6,8-diiodo-7-methoxyflavone (5)
A mixture of 6,8-diiodo-5,7-dihydroxyflavone (1 equiv.)
and K2CO3 and dimethylsulfate (1.1 equiv.) in acetone was
refluxed for 4 h, monitoring by TLC. The mixture was fil-
5.1.1. 6,8-Dibromo-5,7-dihydroxyflavone (2)
To the flask dried on the flame was placed 1.8 ml of Me2S
and immediately dissolved with 20 ml of anhydrous CH2Cl2.

H. Park et al. / European Journal of Medicinal Chemistry 40 (2005) 943–948
947
1
tered and filtration was concentrated. Solidified product was
filtered, washed with water, and recrystallized from metha-
nol to form the titled flavone. 1H-NMR (200 MHz, DMSO-
d6): d 14.06 (s, 1H, OH-5), 8.31–8.35 (m, 2H, H2’ and H6′);
7.31–7.61 (m, 3H, H3’, H4’ and H5’); 7.41 (s, 1H, H3); 3.97
(s, 3H, OCH3). 13C-NMR (50 MHz, DMSO-d6): d 182.1
(C-4 and C-7), 164.4 (C-2), 161.3 (C-5 and 9), 132.8 (C-1’),
130.2 (C-3’ and C-5’), 129.4 (C-4’), 126.4 (C-2’ and C-6’),
107.7 (C-10), 105.3 (C-3), 97.3 (C-6 and C-8), 60.8 (OCH3).
EIMS: m/z 520 (M + 1). Anal. C16H10I2O4 (C, H, N).
obtain a pure product. H-NMR (200 MHz, DMSO-d6): d
13.42 (s, 1H, OH5), 9.45 (s, 1H, OH7); 7.75–7.79 (m, 3H,
phenyl); 7.48–7.58 (m, 12H, phenyl); 7.10 (s, 1H, H3). 13C-
NMR (50 MHz, DMSO-d6): d 182.7 (C-4), 163.2 (C-2), 158.3
(C-7), 157.6 (C-5), 153.2 (C-9), 132.3 (2C-1”), 131.7 (C-1’),
129.1 (2C-3” and 2C-5”), 128.3 (C-3’and C-5’), 127.8 (C-4’),
127.3 (2C-2”, 2C-4” and 2C-6”), 126.2 (C-2’and C-6’), 113.6
(C-6), 109.2 (C-10), 104.8 (C-8), 97.3 (C-3). EIMS: m/z 406
(M+). Anal. C27H18O4 (C, H, N).
5.1.8. 5,7-Dihydroxy-6,8-dimethylsulfinylflavone (9)
5.1.5. 6,8-Diiodo-5,7-dimethoxyflavone (6)
Ferric chloride and dimethyl disulfide were added to a solu-
tion of chrysin in dried toluene with vigorously stirring. The
mixture was then heated to 105 °C and kept at this tempera-
ture for 14 h.After cooling to the room temperature, hydroly-
sis was carried out using solution of 10% hydrochloric acid.
The mixture was extracted with dichloromethane (5 × 15 ml),
and the solvent was evaporated. The solid was crystallized
from methanol to obtain 6,8-dimethylthiochrysin), 87% yield.
1H-NMR (200 MHz, DMSO-d6): d 14.03 (s, 1H, OH), 10.37
(s, 1H, OH), 8.01–8.06 (q, 2H, H2’, H6’), 7.57–7.59 (t, 3H,
H3’, H4’, H5’), 6.78 (s, 1H, H3), 2.39–2.41 (s, 6H, 2 × SCH3).
13C-NMR (50 MHz, DMSO-d6): d 182.3 (C-4), 165.1 (C-2),
163.5 (C-7), 162.7 (C-10), 157.2 (C-9), 132.4 (C-1’), 130.7
(C-3’ and C5’), 129.1 (C-4’), 126.5 (C-2’ and C-6’), 105.1
(C-9), 104.8 (C-6 and C-8), 99.6 (C-3), 18.0 and 16.9 (2 ×
SCH3). EIMS: m/z 346 (M + 1).Anal. C17H14O4S2 (C, H, N).
Methylation of the 5,7-phenolic groups was carried out
following the procedure for the synthesis of the compound 3.
1H-NMR (200 MHz, CDCl3): d 8.04-8.09 (d, 2H, J = 7.4 Hz,
2 Hz, H2’, H6’); 7.54–7.58 (m, 3H, J = 7.2 Hz, 2.2 Hz, H3’,
H4’, H5’); 6.79 (s, 1H, H3); 3.96–3.99 (s, 6H, 2 × OCH3).
13C-NMR (50 MHz, DMSO-d6): d 175.0 (C-7), 163.4 (C-4),
161.0 (C-5), 160.1 (C-2), 157.4 (C-9), 132.1 (C-1’), 130.5
(C-3’ and C-5’), 129.2 (C-4’), 126.5 (C-2’ and C-6’), 115.7
(C-10), 107.8 (C-3), 89.0 (C-8), 79.4 (C-6), 61.5 and 60.7 (2
× OCH3). EIMS: m/z 534 (M+). Anal. C17H12I2O4 (C, H, N).
5.1.6. 5,7-Dimethoxy-6,8-diphenylflavone (7)
To a solution of 5,7-dimethoxy-6,8-diiodoflavone in DME
was added Pd (PPh3)4 (0.2% mole) The resulting mixture was
degassed and stirred at ambient temperature for 20 min. before
the addition Na2CO3 solution. The mixture was degassed
again and then stirred in an atmosphere of nitrogen for sev-
eral hours. The benzeneboronic acid (3 equiv.) was added,
and the reaction mixture was heated at 80 °C for 2–4 h with
monitoring by TLC. After cooling to room temperature, the
mixture was diluted with dichloromethane and water, the
organic phase was separated, washed with water and dried
over magnesium sulfate, filtered and evaporated in vacuum.
The remain solid was purified by flash column to obtain 5,7-
dimethoxy-6,8-diphenylflavone, yield 76%. 1H-NMR
(200 MHz, CDCl3): d 7.36–7.55 (m, 15H, J = 8.0 Hz, 2.8,
H-phenyl); 6.75 (s, 1H, H3); 3.16–3.60 (s, 6H, 2 × OCH3).
13C-NMR (50 MHz, DMSO-d6): d 176.3 (C-4), 160.5 (C-2),
132.2 (C-7), 131.8 (C-5), 131.5(C-9), 130.7 (2C-1”, 130.4
(C-1’), 128.9 (2C-3” and 2C-5”), 128.3 (2C-3”), 127.9 (2C-
2”, 2C-4” and 2C-6”), 125.9 (C-2’ and C-4’), 124.3 (C-6),
121.2 (C-10), 108.6 (C-3), 61.8 and 60.7 (2 × OCH3). EIMS:
m/z 434 (M + 1). Anal. C29H22O4 (C, H, N).
5.1.9. 5,7-Dimethoxy-6,8-dimethylsulfinylflavone (10)
Methylation of the compound 9 was carried out following
the procedure for the synthesis of the compound 3. 1H-NMR
(200 MHz, DMSO-d6): d 8.02–8.04 (q, 2H, H2’, H6’); 7.54–
7.59 (t, 3H, H3’, H4’, H5’); 6.80 (s, 1H, H3); 4.03 and 4.07
(s, 6H, 2-OCH3); 2.51 and 2.47 (s, 6H, 2 × SCH3). 13C-NMR
(50 MHz, DMSO-d6): d 176.5 (C-4), 165.7 (C-2), 162.3 (C-7),
160.9 (C-5), 157.3 (C-9), 131.1 (C-1’), 128.7 (C-3’and C5’),
125.9 (C-4’), 125.6 (C-2’ and C-6’), 108.4 (C-10), 107.8
(C-8), 104.9 (C-6), 95.4 (C-3), 61.3 (OCH3), 61.0 (OCH3),
17.8 (2 × SCH3). EIMS: m/z 374 (M+). Anal. C19H18O4S2
(C, H, N).
5.1.10. 5,7-Dihydroxy-6,8-dimethoxyflavone (11)
5,7-Dihydroxyflavone (chrysin) was protected using ben-
zyl bromide (1 equiv.) and K2CO3 in the refluxing acetone
solvent. Then, the benzyl protected flavone (1a) was treated
with excess potassium persulfate (3 equiv.) in the present of
potassium hydroxide and pyridine to afford 7-benzoyl-5,6,8-
trihydroxyflavone (1b). This product was methylated with
dimethyl sulfate (2 equiv.) to obtain 7-benzoyl-6,8-dimethoxy-
5-hydroxyflavone (1c), followed by hydrolysis to obtain the
5.1.7. 5,7-Dihydroxy-6,8-diphenylflavone (8)
Solution 1 M BBr3 in CH2Cl2 (4 equiv.) was added slowly
to the solution of 57-Dimethoxy-6,8-diphenylflavone during
10 min. Nitrogen gas was inserted to the reaction flask to
remove the oxygen gas, then it was heating at 40 °C for 6–10 h,
monitoring by TLC. The reaction mixture was cooled to room
temperature, methanol was added to destroy the excess of
BBr3. Methanol and solvent was removed in vacuum to obtain
solid. The solid was washed with water and recrystallized
twice from acetone-water or dichloromethane - methanol to
1
titled product. H-NMR (200 MHz, DMSO-d6): d 12.78 (s,
1H, OH), 10.64 (s, 1H, OH), 8.15–8.19 (q, 2H, H2’, H6’),
7.68–7.73 (t, 3H, H3’, H4’, H5’), 7.12 (s, 1H, H3), 3.88–3.98
(s, 6H, 2 × OCH3). 13C-NMR (50 MHz, DMSO-d6): d 182.5
(C-4), 163.1 (C-2), 148.4 (C-5), 147.7 (C-10), 138.7 (C-7),
131.4 (C-1’), 130.6 (C-6), 130.2 (C-8), 128.6 (C-3’and C-5’),

948
H. Park et al. / European Journal of Medicinal Chemistry 40 (2005) 943–948
125.7 (C-2’, C-4’and C-6’), 104.6 (C-9), 97.2 (C-3), 61.2 and
60.4 (2 × OCH3). EIMS: m/z 314 (M+). Anal. C17H14O6 (C,
H, N).
search Program of the Korea Science and Engineering Foun-
dation. The authors would like to thank Pharmacal Research
Institute and Central Laboratory of Kangwon National Uni-
versity for the use of analytical instruments and bioassay
facilities.
5.1.11. 5,6,7,8-Tetrahydroxyflavone (12)
Debenzylation of 7-benzoyl-5,6,8-trihydroxyflavone (1b)
in aqueous c-HCl solution gave the titled compound 1H-NMR
(200 MHz, DMSO-d6): d 12.27 (s, 1H, OH), 10.07 (s, 1H,
OH); 9.04 (s, 2H, OH); 8.25–8.27 (m, 2H, H2’, H6’); 7.67–
7.71 (t, 3H, H3’, H4’, H5’); 7.01 (s, 1H, H3). 13C-NMR
(50 MHz, DMSO-d6): d 182.6 (C-4), 162.7 (C-2), 142.9 (C-9),
139.8 (C-5), 139.1 (C-7), 131.8 (C-1’), 129.2 (C-3’and C5’),
126.5 (CC4’ and C6), 126.4 (C-8), 125.6 (C-2’ and C-6’),
104.1 (C-10), 103.0 (C-3). EIMS: m/z 286 (M+). Anal.
C15H10O6 (C, H, N).
References
[1] M. Hecker, C. Preiss, P. Klemm, R. Brusse, Br. J. Pharmacol. 118
(1996) 2178–2184.
[2] F.L. Pearce, J. Bienenstock, J. Allergy Clin. Immunol. 73 (1984)
819–824.
[3] R.J. Fishkin, J.T. Winslow, Psychopharmacology (Berl.) 132 (1997)
335–341.
[4] S. Habtemariam, J. Nat. Prod. 60 (1997) 775–778.
5.1.12. 5,6,7,8-Tetramethoxyflavone (13)
[5] Y.C. Liang, S.H. Tsai, D.C. Tsai, S.Y. Lin-Shiau, J.K. Lin, FEBS Lett.
496 (2001) 12–18.
A mixture of 5,6,7,8-tetrahydroxyflavone (1 equiv.) and
K2CO3 and dimethylsulfate (5.2 equiv.) in acetone was
refluxed for overnight, monitoring by TLC. The mixture was
filtered and filtration was concentrated. Solidified product was
filtered, washed with water, and recrystallized from metha-
nol to form the titled flavone. 1H-NMR (200 MHz, CDCl3): d
7.90–7.95 (m, 2H, H2’and H6’); 7.43–7.58 (m, 3H, H3’, H4’
and H5’); 3.95, 3.98, 4.03 and 4.11 (s, 12H, 4 × OCH3). 13C-
NMR (50 MHz, CDCl3): d 176.8 (C-4), 161.8 (C-2), 146.5
(C-7), 138.8 (C-9), 130.9 (C-1’), 128.5 (C-3’and C5’, C-6 and
C-8), 125.4 (C-2’and C-6’), 107.4 (C-10), 95.7 (C-3), 61.07,
61.2, 61.4, 61.6 (4 × OCH3). EIMS: m/z 342 (M+). Anal.
C19H18O6 (C, H, N).
[6] Y.-C. Chen, L.-L. Yang, T.J.-F. Lee, Biochem. Pharmacol. 59 (2000)
1445–1457.
[7] Y.S. Chi, B.S. Cheon, H.P. Kim, Biochem. Pharmacol. 61 (2001)
1195–1203.
[8] T.T. Dao, Y.S. Chi, J. Kim, H.P. Kim, S. Kim, H. Park, Arch. Pharm.
Res. 26 (2003) 345–350.
[9] T.T. Dao, J.W. Oh, Y.S. Chi, H.P. Kim, K.-S. Sin, H. Park, Arch.
Pharm. Res. 26 (2003) 581–584.
[10] B. Samuel, US Patent 4,517,388 (1985).
[11] R.H. de Rossi, A.V. Veglia, Tetrahedron Lett. 27 (1986) 5963–5966.
[12] A. Suzuki, J. Organomet. Chem. 576 (1996) 147–168.
[13] V. Catherine, L. Yves, P. Jean-Marie, E. Aquitaine, US Patent
5,113,019 (1992).
[14] E.J. Behrman, in:A.S. Kende (Ed.), Organic Reactions, vol. 35, Wiley,
New York, Chichester, Brisbane, Toronto, Singapore, 1988, pp. 421–
492.
Acknowledgements
This work was supported by the grant (R05-2003-000-
11789-0 and R01-2004-000-10134-0) from the Basic Re-
[15] T. Mossman, J. Immnol. Methods 65 (1983) 55–63.