Synthesis of licochalcone analogues with increased anti-inflammatory activity

Article abstract of DOI:10.1016/j.bmcl.2013.11.044

Licohalcones have been reported to have various biological activities. However, most of licochalcones also showed cytotoxicity even though their versitile utilities. Licochalcones B and D, which have common substituents at aromatic ring B, are targeted to

Full text of DOI:10.1016/j.bmcl.2013.11.044

Bioorganic & Medicinal Chemistry Letters 24 (2014) 181–185
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Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis of licochalcone analogues with increased
anti-inflammatory activity
Si-Jun Kim a, Cheol Gi Kim a, So-Ra Yun a, Jin-Kyung Kim b, Jong-Gab Jun a,
⇑
a Department of Chemistry and Institute of Applied Chemistry, Hallym University, Chuncheon 200-702, Republic of Korea
b Department of Biomedical Science, College of Natural Science, Catholic University of Daegu, Gyeungsan-Si 700-702, Republic of Korea
a r t i c l e i n f o
a b s t r a c t
Article history:
Licohalcones have been reported to have various biological activities. However, most of licochalcones also
showed cytotoxicity even though their versitile utilities. Licochalcones B and D, which have common sub-
stituents at aromatic ring B, are targeted to modify the structure at aromatic ring A for inflammatory
studies. Licochalcone derivatives (1–6) thus prepared are compared for their suppression ability of nitric
Received 1 October 2013
Revised 31 October 2013
Accepted 18 November 2013
Available online 26 November 2013
oxide (NO) production and showed 9.94, 4.72, 10.1, 4.85, 2.37 and 4.95 lM of IC50 values, respectively.
Ó 2013 Elsevier Ltd. All rights reserved.
Keywords:
Licohalcone B
Licohalcone D
[3,3]-Sigmatropic rearrangement
anti-Inflammatory effects
IC50 values of NO production
Licochalcone A (LicoA), licochalcone B (LicoB), licochalcone C
(LicoC), licochalcone D (LicoD), echinatin and lsoliquiritigenin are
major active components in licorice which is a traditional medicine
used in the Northeast Asia for the treatment of ulcer, asthma,
inflammation and other diseases.1 Licochalcones have been iso-
lated and characterized from the root of Glycyrrhiza inflata, and
have been reported to show various biological properties, includ-
ing antibacterial,2 antitumor,3 anti-inflammatory,4 and antioxida-
tive5 activities. G. inflata is the main species in licorice and
contains about 40 kinds of flavonoids of which having no hydroxyl
at the position 20 (or 60) which is different from usual flavonoid
have been found to show the major contribution on biological
activities.6 These unusual chalcones are called retrochalcone or ‘re-
versely constructed chalcone’ in which the ring A would be derived
from shikimate and the ring B from polyketide of malonate orgin.7
The anti-inflammatory activities of licochalcones as shown in
Scheme 1 were compared by the inhibition effect of the mast cell
degranulation which play a key role in allergic inflammation in
RBL (rat basophilic leukemia)-2H3 cells.8 Also, the anti-inflamma-
tory activities were compared by the inhibition effect of NO
production in inflammatory regions.9 The 50% inhibitory concen-
tration (IC50) against degranulation, 30% cytotoxicity (CC30), and
IC50 of LPS-induced NO production of each licochalcones are listed
in Scheme 1. Since LicoA, LicoC and LicoD exhibited similar inhib-
itory effects on the degranulation with the IC50 at 17, 24 and
21
l
M. Also, LicoB and LicoD showed IC50 of LPS-induced NO pro-
duction at 2.3 and 2.2 lM, respectively. From these inhibitory re-
sults, we found that LicoB and LicoD having common
substituents at aromatic ring B showed higher activity with rela-
tively lower cytotoxicity than the others.
The structure of LicoB is similar to echinatin except the pres-
ence or absence of 3-hydroxy substituent at ring B, however, the
anti-inflammatory activities are quite different and only LicoB
showed the inhibition activity of 2.3 lM for IC50 of NO production,
but both compounds showed lower cytotoxicity. Also the structure
of LicoB is exactly same with LicoD on ring B which has 3,4-dihy-
droxy-2-methoxy substituents, and both showed the higher inhibi-
tion activities for IC50 of NO production. In order to find a highly
active anti-inflammatory licochalcone derivative having lower
cytotoxicity, we designed the structures same as LicoB and LicoD
at ring B, which have 3,4-dihydroxy-2-methoxy substituents, but
modified at ring A to resorcinol type (2), catechol type (3) and rear-
ranged isopropenyl analogues 5 and 6 (Scheme 2).
The natural LicoB (1) was first identified in 1975 from the roots
of Glycyrrhiza glabra Linn. (licorice from Sinkiang, China)7 and has
been reported many biological activities,8–10 however, only single
report for synthesis has been known.11 Recently, we reported the
first total synthesis of LicoD12 and we used the similar route for
the synthesis of LicoB as shown in Scheme 3 since the ring B of
LicoB is exactly same as LicoD. The aldehyde portion (9) of pro-
tected ring B is condensed with THP protected acetophenone (11)
using 3 M NaOH in MeOH to produce the chalcone (12), and fol-
lowing deprotection using Dowex 50X2 resin afforded the LicoB
⇑
Corresponding author. Tel.: +82 33 248 2075; fax: +82 33 256 3421.
E-mail address: jgjun@hallym.ac.kr (J.-G. Jun).
0960-894X/$ - see front matter Ó 2013 Elsevier Ltd. All rights reserved.
http://dx.doi.org/10.1016/j.bmcl.2013.11.044

182
S.-J. Kim et al. / Bioorg. Med. Chem. Lett. 24 (2014) 181–185
O
O
O
b
6
B
3
6'
A
3'
1
2
5'
4'
1'
2'
5
a
OH
MeO
MeO
HO
OH
HO
HO
MeO
OH
4
OH
LicoA
LicoB
LicoC
>30
>30
2.3
24
>30
-
IC50 against degranulation (µM) 17
CC30 (µM)
24
IC50 of NO production (µM)
5.6
O
OH
O
O
MeO
OH
HO
OH
HO
MeO
OH
HO
OH
Isoliquiritigenin
LicoD
Echinatin
>30
>30
-
21
>30
2.2
>30
>30
-
Scheme 1. anti-Inflammatory activity of licochalcones.
O
O
O
OH
HO
HO
MeO
OH
HO
MeO
OH
MeO
OH
HO
OH
OH
OH
LicoB (1)
3
2
O
O
O
OH
OH
MeO
MeO
HO
OH
HO
MeO
HO
OH
OH
OH
5
6
LicoD (4)
Scheme 2. The designed structures of licochalcone B and D derivatives.
in total 25% yield of 5 steps. The spectral data for LicoB agreed well
with the literature values.7,10,11
1-bromo-3-methyl-2-butene in K2CO3 to give 21 in 98% yield,
which is then condensed with the aldehyde 9 to give chalcone 22
in 86% yield, and following water-accelerated Claisen rearrange-
ment14 produced the analogue 5 in 52% yield (Scheme 6).
Another isopropenyl analogue 6, which is also a new compound,
is prepared from 10 using same methododlogy via O-prenylated
acetophenone 23 and chalcone 24 as shown in Scheme 7.
Resorcinol (2) analogue of LicoB, which is a new compound, was
prepared by conventional Claisen–Schmidt condensation of 2,4-
diethoxymethoxyacetophenone (14) with the benzaldehyde (9)
using basic condition followed by deprotection in moderate yields
(Scheme 4).
3,4-Diethoxymethoxyacetophenone (19) was prepared from
commercially available 3,4-dihydroxybenzaldehyde (16) by pro-
tection using diisopropylethylamine with EOMCl followed by CH3-
MgCl reaction and PDC oxidation (Scheme 5). It is noteworthy that
the deprotonation of 2-hydroxyacetophenone (13) requires stron-
ger base than 3-hydroxy analogue (16) because of intramolecular
hydrogen bonding. The acetophenone 19 afforded the catechol
analogue (3), which is a known natural licochalcone as tetrahydr-
oxymethoxychalcone,13 by condensation and deprotection.
Compound 5, a new compound, is an anologue of LicoD, which
has a similar isopropenyl substituent to LicoA. [3,3]-Sigmatropic
rearrangement in LicoD synthesis has been applied for this synthe-
sis.12 4-Hydroxyacetophenone (10) is O-prenylated using
The synthetic licochalcones 1–6 are subjected to comparison of
their anti-inflammatory activities. The suppression results of nitric
oxide (NO) production in lipopolysaccharide (LPS)-stimulated
RAW264.7 machrophages for the licochalcones 1–6 are compared
in Table 1. Isopropenyl analogue 2 and resorcinol analogue 5
showed 75.9% and 71.1% suppression of NO production at 2 lM,
respectively. LicoD (4) and its analogue 6 also showed moderate%
inhibition at same concentration, however, LicoB (1) and its cate-
chol analogue 3 showed very low or no inhibition activities. Also,
Resorcinol 2, isopropenyl 5, LicoD (4) and its analogue 6 showed
96.5%, 96.1%, 95.4% and 94.3% inhibition, respectively, at 20 lM.
Interestingly, catechol analogue showed 92.5% inhibition, however,
LicoB (1) still showed only 69.6% inhibition at this concentration.

S.-J. Kim et al. / Bioorg. Med. Chem. Lett. 24 (2014) 181–185
183
O
O
O
K2CO3, CH3I
iPr2NEt, EOMCl
H
H
H
CH2Cl2, rt
12h, 58%
acetone, reflux
3.5h, 90%
MeO
OMOE
OMOE
HO
OH
HO
OMOE
OMOE
OH
9
7
8
O
3M NaOH
Dowex 50X2 resin
LicoB
MeOH, rt
6h, 64%
MeOH, rt, 12h
84%
THPO
MeO
OMOE
OMOE
12
O
O
DHP, Py-pTsOH
CH2Cl2, rt
3h, 89%
OH
OTHP
10
11
Scheme 3. Synthesis of licochalcone B (1).
O
O
O
EOMO
9
K2CO3, EOMCl
DMF, 70oC
KOH, EtOH, rt
EOMO
MeO
OMOE
OMOE
OH
HO
14h, 71%
OMOE
EOMO
8h, 45%
13
14
15
Dowex 50X2 resin
2
MeOH, rt, 3h
75%
Scheme 4. Resorcinol analogue (2) of licochalcone B.
O
O
OH
OH
OH
iPr2NEt, EOMCl
CH3MgCl
THF,rt
1h, 98%
OMOE
OMOE
OMOE
H
H
CH2Cl2, rt
20h, 83%
OMOE
16
18
17
O
O
EOMO
EOMO
PDC
9
OMOE
OMOE
CH2Cl2, rt
9h, 96%
MeO
OMOE
OMOE
KOH, EtOH, rt
14h, 71%
20
19
Dowex 50X2 resin
3
MeOH, rt, 15h
40%
Scheme 5. Catechol analogue (3) of licochalcone B.

184
S.-J. Kim et al. / Bioorg. Med. Chem. Lett. 24 (2014) 181–185
O
O
9
K2CO3
Br
10
KOH, EtOH, rt
18h, 86%
acetone, reflux
1.5h, 98%
O
MeO
OMOE
OMOE
O
22
21
Bomb reactor
EtOH/H2O (v/v)=4/1
5
150oC, 20h
52%
Scheme 6. Synthesis of licochalcone analogue 5.
O
O
9
Br
K2CO3
10
KOH, EtOH, rt
36h, 87%
acetone,reflux
4h, 99%
O
MeO
OMOE
OMOE
O
24
23
Bomb reactor
EtOH/H2O (v/v)=4/1
6
150oC, 24h
51%
Scheme 7. Synthesis of licochalcone analogue 6.
Table 1
anti-Inflammatory activities of licochalcones 1–6
Compound
NO production (% inhibition)
20
5.4 0.3 (94.6)
2
l
M
lM
Medium (MED)
1
2
3
4
5
6
LPS
5.4 0.3 (94.6)
82.8 0.3 (17.2)
28.9 0.3 (71.1)
30.4 0.3 (69.6)
3.5 0.3 (96.5)
7.5 0.8 (92.5)
4.6 0.2 (95.4)
3.9 0.1 (96.1)
5.7 0.3 (94.3)
100.0 0.8 (0.0)
106.0 6.0 (ꢀ6.0)
42.2 0.6 (57.8)
24.1 1.0 (75.9)
43.5 0.6 (56.5)
100.0 0.8 (0.0)
The results are reported as mean value SEM for n = 3.
% Inhibition is based on LPS as shown in parenthesis.
Table 2
Proliferation effect of licochalcones 1–6
MED
1
2
3
4
5
6
2
l
M
1.9
1.9
2.0
2.1
2.3
2.3
2.0
2.4
1.8
0.5
1.8
0.6
1.9
0.6
20 lM
Figure 1. IC50 values for NO production of synthetic licochalcones 1–6.
The cell viability assay at 20 lM concentration was not affected by
activation. Based on these data, inhibitory effect of NO production
may due to the blocking of NF-jB activation.
In summary, we prepared licochalcone analogues 1–6 by con-
ventional Claisen–Schmidt condensation in basic condition. In
anti-inflammatory studies, all the compounds tested (1–6) showed
69.6%, 96.5%, 92.5%, 95.4%, 96.1% and 94.3% suppression of NO pro-
the licochalcones 1–3 indicating no cytotoxicity as shown in
Table 2. LicoD (4) and its isopropenyl analogues 5–6, unfortunately,
showed significant cytotoxicities, and the inhibition activities of
4–6 considered due to cytotoxicities. IC50 values for NO production
of synthetic licochalcones 1–6 were evaluated by using GraphPad
Prism 4.0 software and showed 9.94, 4.72, 10.1, 4.85, 2.37 and
duction, respectively, at 20
4.85, 2.37 and 4.95 M of IC50 values, respectively. However, LicoD
(4) and its analogues 5–6 showed significant cytotoxicities, but the
lM and also showed 9.94, 4.72, 10.1,
4.95
Previously reported data15–17 showed that various licochalcones
significantly inhibited LPS-induced nuclear factor (NF)-
lM, respectively (Fig. 1).
l
jB

S.-J. Kim et al. / Bioorg. Med. Chem. Lett. 24 (2014) 181–185
185
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This research was financially supported by the Ministry of
Education, Science Technology (MEST) and Korea Institute for
Advancement of Technology (KIAT) through the Human Resource
Training Project for Regional Innovation (2012H1B8A2026036),
and by Priority Research Centers Program through the National
Research Foundation of Korea (NRF) funded by the Ministry of
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Supplementary data
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2013.11.044.
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