Synthesis and biological evaluation of 2-aroylbenzofurans, rugchalcones A, B and their derivatives as potent anti-inflammatory agents

Article abstract of DOI:10.1016/j.bmcl.2016.02.023

An efficient synthesis of 2-aroylbenzofurans, rugchalcones A, B and their derivatives was accomplished in excellent yields by the Rap–Stoermer reaction between substituted salicylaldehydes and phenacyl bromides. Later their anti-inflammatory effects were evaluated in lipopolysaccharide (LPS)-induced RAW-264.7 macrophages. The compounds were exhibited exceptional potency against inflammatory mediated NO production with no cytotoxicity at 10?μM concentration and IC₅₀values are found in the range from 0.75 to 13.27?μM. Among the 2-aroylbenzofurans prepared in this study, compounds 4 (99.6%; IC₅₀?=?0.57), rugchalcone B (2) (99.3%; IC₅₀?=?4.13), 7 (96.8%; IC₅₀?=?1.90) and 8 (74.3%; IC₅₀?=?0.99) were showed the maximum inhibitory activity. This study suggests that compounds 2, 4, 7 and 8 which are having 4-hydroxyphenyl group and/or hydroxy (–OH) group at 5- and/or 6-position of benzofuran motif could be considered as a promising scaffolds for the further development of iNOS inhibitors for potential anti-inflammatory applications.

Full text of DOI:10.1016/j.bmcl.2016.02.023

Bioorganic & Medicinal Chemistry Letters 26 (2016) 1521–1524
Contents lists available at ScienceDirect
Bioorganic & Medicinal Chemistry Letters
journal homepage: www.elsevier.com/locate/bmcl
Synthesis and biological evaluation of 2-aroylbenzofurans,
rugchalcones A, B and their derivatives as potent anti-inflammatory
agents
Young Hwa Seo a, Kongara Damodar a, Jin-Kyung Kim b, Jong-Gab Jun a,
⇑
a Department of Chemistry and Institute of Natural Medicine, 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:
An efficient synthesis of 2-aroylbenzofurans, rugchalcones A, B and their derivatives was accomplished in
excellent yields by the Rap–Stoermer reaction between substituted salicylaldehydes and phenacyl bro-
mides. Later their anti-inflammatory effects were evaluated in lipopolysaccharide (LPS)-induced RAW-
264.7 macrophages. The compounds were exhibited exceptional potency against inflammatory mediated
Received 24 October 2015
Revised 28 January 2016
Accepted 10 February 2016
Available online 10 February 2016
NO production with no cytotoxicity at 10
lM concentration and IC50 values are found in the range from
0.75 to 13.27 M. Among the 2-aroylbenzofurans prepared in this study, compounds
l
4
(99.6%;
Keywords:
2-Aroylbenzofuran
Rugchalcones
Rap–Stoermer reaction
Nitric oxide
anti-Inflammatory
IC50 = 0.57), rugchalcone B (2) (99.3%; IC50 = 4.13), 7 (96.8%; IC50 = 1.90) and 8 (74.3%; IC50 = 0.99) were
showed the maximum inhibitory activity. This study suggests that compounds 2, 4, 7 and 8 which are
having 4-hydroxyphenyl group and/or hydroxy (–OH) group at 5- and/or 6-position of benzofuran motif
could be considered as a promising scaffolds for the further development of iNOS inhibitors for potential
anti-inflammatory applications.
Ó 2016 Elsevier Ltd. All rights reserved.
In multi-cellular organisms, inflammation is a cardinal host
defense response to tissue damage, injury, infectious agents or
autoimmune responses and is an integral part of the immune
response.1 Symptoms of inflammation include swelling, redness
of the area, pain, and sometimes loss of function.2 Based on time
and pathological features, it can be either acute or chronic. Inflam-
mation is present in several disorders and diseases like atheroscle-
rosis, diabetes and cancer. Increased blood supply, enhanced
vascular permeability and migration of immune cells occur at
damaged sites. In this process, activated inflammatory cells (neu-
trophils, eosinophils, mononuclear phagocytes and macrophages)
secrete increased amounts of nitric oxide (NO), prostaglandins
(PGs) and cytokines, such as interleukin (IL)-1b, IL-6, and tumor
necrosis factor (TNF). Among these, two of the most prominent
are PGs and NO. PGs are produced by cyclooxygenase (COX, which
mainly having two forms COX-1 and COX-2) by arachidonic acid
pathway. NO is a small, lipophilic, diffusible and transient free-rad-
including vasodilatation, thermoregulation, and neuromodulation.
High levels of NO is produced ‘on-demand’ by the inducible (iNOS)
enzyme, to help kill tumors, viruses and bacteria. Both underpro-
duction and overproduction of NO have been linked to various
human pathologies. Insufficient NO production from eNOS and
nNOS can lead to hypertension, atherosclerosis, and cardiovascular
disease, whereas excess NO production by iNOS can cause inflam-
mation, inflammatory bowel disease (IBD), rheumatoid arthritis,
asthma, diabetes, stroke, cancer and neurodegenerative disorders.3
Therefore, control of the excess NO production by inhibition of
iNOS may exert anti-inflammatory effects.
Traditional non-steroidal anti-inflammatory drugs (tNSAIDs)
and aspirin usage is general practice in the therapeutic approach
to alleviate the symptoms associated with both acute and chronic
inflammatory diseases. Their activity is most likely mediated
through their ability to inhibit COX enzymes. However, their
long-term oral administration is restricted because of the high inci-
dence of side effects, particularly those relating to the gastroin-
testinal (GI) tract, renal and cardiovascular systems due to the
inhibition of the housekeeping enzyme COX-1 along with COX-
2.4 Later, selective COX-2 inhibitors (COXIBs) were introduced to
reduce the risks. While these COXIBs did reduce the risk of GI
injury, like the tNSAIDs, they are also appeared to increase the risk
of cardiovascular events, such as heart attack and stroke. Hence,
ical species generated from L-arginine by three types of nitric oxide
synthase (NOS) enzymes. It acts as a double-edged sword. Physio-
logically vital amount of NO produced by the endothelial (eNOS)
and neuronal (nNOS) enzymes which is crucial for signaling,
⇑
Corresponding author. Tel.: +82 33 248 2075; fax: +82 33 256 3421.
E-mail address: jgjun@hallym.ac.kr (J.-G. Jun).
http://dx.doi.org/10.1016/j.bmcl.2016.02.023
0960-894X/Ó 2016 Elsevier Ltd. All rights reserved.

1522
Y. H. Seo et al. / Bioorg. Med. Chem. Lett. 26 (2016) 1521–1524
there remains an appeal to search for more effective anti-inflam-
matory drugs with minimal side-effects.
sensors and organic semiconductors.14 Their wide range of phar-
macological and physical properties created special interest to
researchers towards the design and synthesis of these important
heterocyclic compounds.
Rugchalcones A and B (Fig. 1) are 2-aroylbenzofuran derivatives
isolated from the flowers of Rosa rugosa and displayed anti-tobacco
mosaic virus (anti-TMV) activities.15 In continuation of our
work16 on the synthesis of bioactive natural products and their
analogs, herein we wish to describe an efficient synthesis and
anti-inflammatory activity evaluation of rugchalcones A (1), B (2)
and their derivatives (3–8).
Benzofurans and their derivatives in particular are important
scaffolds for drug development.5 Several natural and non-natural
2-substituted benzofurans have been noted for their antioxidant,6
antifungal,7 antimicrobial,8 anti-inflammatory,9 PPAR-d agonists,10
antitubercular,11 anti-HIV, anti-tumor and anti-platelet activity.12
Radiolabeled benzofuran derivatives were used as molecular imag-
ing probes for b-amyloid plaques in Alzheimer’s Disease (AD).13
Besides this, few derivatives were find application as fluorescent
Our approach for the synthesis of rugchalcones A (1), B (2) and
their derivatives (3–8) is outlined in Scheme 1. The synthesis com-
menced with the preparation of 2,4,5-trihydroxybezaldehyde (10)
from 1,2,4-benzenetriol (9). Regioselective methylation of phenolic
4-OH group of 10 was carried out using equimolar amounts of
methyl iodide (MeI) and KOH in N,N-dimethylformamide (DMF)
to afford compound 11 in 64% yield which was subsequently pro-
tected using tert-butyldimethylsilyl chloride (TBDMSCl) to furnish
substituted salicylaldehyde, 12 in 86% yield. While, phenolic 4-OH
group of 10 regioselectively protected with ethoxymethyl (EOM)
group using chloromethyl ethyl ether (EOM-Cl), triethylamine
(Et3N) and catalytic tetrabutylammonium iodide (TBAI) in acetone
and we were pleased to isolate compound 13 in 96% yield which
O
10
1'
4
7
R1
R2
3
9
8
2'
6
2
4'
O
R3
5'
R1 = OH, R2, R3 = OMe; Rugchalcone A (1)
Rugchalcone B
5
R1, R2, R3 = OMe; ( )
R1, R3 = OH, R2 = OMe;
6
R1, R2 = OMe, R3 =OH; ( )
2
( )
R1, R3 = OMe, R2 = OH; ( )
7
R1, R2 = OH, R3 =OMe; ( )
3
R1, = OMe, R2, R3 = OH; ( )
R1, R2, R3 = OH; ( )
4
8
Figure 1. Structures of rugchalcones A (1), B (2) and their derivatives (3–8).
HO
HO
HO
HO
CHO
OH
HO
CHO
OH
TBSO
MeO
CHO
OH
a
c
b
64%
52%
86%
OH
MeO
10
HO
11
MeO
9
12
HO
HO
CHO
OH
CHO
OH
CHO
OH
d
96%
e
54%
MOEO
MOEO
14
10
13
15
HO
HO
CHO
OH
MOEO
MOEO
CHO
OH
MeO
MeO
CHO
OMe
MeO
MeO
CHO
f
g
98%
64%
OH
16
17
10
O
O
O
O
Br
h
98%
i
86-88%
R
HO
TBSO
R
18
19
20
19
R = OTBS
R = OMe
21
22
R = OTBS
R = OMe
O
O
O
R1
R2
R1
R2
R1
R2
CHO
k
Br
j
71-81%
62-99%
O
O
OH
1 = OTBS, R2 = OMe
1 = OMe, R2 = OEOM
1
R
R
R
1
1
1
2
= OH, R2, R = OMe
R , R = OH, R2 = OMe
R , R = OMe, R = OH
1
1
3
4
7
12
22
21
R
R
R = OMe
2
R
= OMe, R2, R = OH
14
15
17
R
R = OTBS
R , R2 = OH, R = OMe
1
R , R2 = OEOM
R , R2 = OMe
R , R = OMe, R2 = OEOM
1
23
24
1
R
1
1 = OMe, R2 = OEOM, R = OH
2
5
6
R , R , R = OMe
R , R2 = OMe, R = OH
1
1
R , R2 = OEOM, R = OMe
25
O
O
MeO
MeO
HO
HO
l
O
O
71%
OMe
OH
5
8
Scheme 1. Reagents and conditions: (a) triethyl orthoformate, AlCl3, benzene, rt, 1 h; (b) MeI, KOH, anhyd DMF, 0–40 °C, 8 h; (c) TBDMSCl, imidazole, DMF, 40 °C, 2.5 h;
(d) chloromethyl ethyl ether, Et3N, TBAI, acetone, 0 °C–rt, 4 h; (e) dimethyl sulfate, K2CO3, acetone, 0 °C–rt, 12 h; (f) chloromethyl ethyl ether, K2CO3, TBAI, acetone, 0 °C–rt,
2.5 h; (g) BCl3, CH2Cl2, À78 °C to rt, 12 h; (h) TBDMSCl, imidazole, DMF, 40 °C, 12 h; (i) CuBr2, EtOAc, reflux, 2 h; (j) K2CO3, acetonitrile, reflux, 1–2 h; (k) DowexÒ resin, MeOH,
THF, rt, 2–4 days; (l) BBr3, CH2Cl2, 0 °C–rt, 28 h.

Y. H. Seo et al. / Bioorg. Med. Chem. Lett. 26 (2016) 1521–1524
1523
upon methylation of 5-OH group gave compound 14. Substituted
salicylaldehyde 15 was also obtained from compound 10 by the
two phenolic–OH groups protection using EOM-Cl, catalytic TBAI
and K2CO3 as base in acetone. Next, 4,5-dimethoxysalicylaldehyde
(17) was obtained in excellent yield from 2,4,5-trimethoxyben-
zaldehyde by demethylation using BCl3.
With the substituted salicylaldehydes 12, 14, 15 and 17 in hand,
our attention shifted to preparation of required phenacyl bro-
mides. Protection of 4-hydroxyacetophenone (18) with tert-
butyldimethylsilyl (TBS) group gave compound 19 in 98% yield.
Next,
a-bromination of compound 19 and 4-methoxyacetophe-
none (20) was carried out using copper(II) bromide (CuBr2) in ethyl
acetate and the corresponding phenacyl bromides 21 and 22 were
obtained in good yields.
We were now ready to subject the Rap–Stoermer reaction17
between substituted salicylaldehydes (12, 14, 15 and 17) and phe-
nacyl bromides (21 and 22), a prominent synthetic protocol
employed for the synthesis of 2-aroylbenzofurans. Treatment of
1.0 equiv of phenacyl bromide, 1.1 equiv of substituted salicylalde-
hyde and 1.3 equiv of K2CO3 in acetonitrile under reflux condition
for 1–2 h led to the isolation of 2-aroylbenzofurans 1, 2, 5, 6 and
23–25 in moderate to excellent yields. Compounds 23–25 were
further subjected to EOM-group deprotection with DowexÒ resin
and we were delighted to obtain the desired rugchalcone deriva-
tives 3, 4 and 7 in 71–81% yields. Demethylation of 5 using excess
BBr3 in CH2Cl2 afforded the product 8 in good yield. The structures
of all the 2-aroylbenzofurans were settled from their spectral (1H
and 13C NMR and MS) data (see the Supplementary data).
In order to evaluate the anti-inflammatory effects of the pre-
pared rugchalcones A (1), B (2) and their derivatives (3–8), we
measured the amount of nitric oxide (NO) which is one of the
essential mediators of inflammation, in lipopolysaccharide (LPS)-
stimulated RAW264.7 macrophages.18
Figure 2. Inhibition of iNOS mediated NO production by compounds 1–8.
Table 2
Proliferation effect of rugchalcones A (1), B (2) and their derivatives (3–8)
Compound
Proliferationa
10 lM
IC50 (lM)
1
l
M
Medium (MED)
100.1 6.7
98.1 5.2
97.9 4.6
93.2 3.3
98.1 2.1
105.4 5.7
105.7 2.0
95.1 3.9
106.3 3.1
98.6 2.9
100.1 6.7
100.1 3.9
93.1 1.7
91.8 3.6
95.9 2.1
98.6 2.6
101.7 1.8
101.2 1.5
100.3 3.3
97.6 5.6
1
2
3
4
5
6
7
8
10.79
4.13
13.27
0.57
11.84
10.42
1.90
0.99
2.69
L
-NMMA
a
The results are reported as mean value SEM for n = 3.
anti-Inflammatory activity: Effect of compounds 1–8 on NO
generation by induced macrophages was monitored (Table 1).
Lipopolysaccharide (LPS) treated RAW 264.7 has been used to
stimulate the production of NO through the activation of iNOS
and NG-monomethyl- -NMMA)19 was employed
L-arginine acetate (L
as positive control. Of the 2-aroylbenzofurans prepared in the pre-
sent study, four compounds, that is, rugchalcone B, compounds 4, 7
and 8 showed significant activities at 10 lM (Fig. 2). Among the 8
compounds, the maximum inhibitory activity was observed with
compound 4 (99.6%) followed by rugchalcone B (99.3%), com-
pounds 7 (96.8%) and 8 (74.3%). The cell viability assay at 10 lM
Figure 3. Effects of compounds 1–8 on iNOS expression (Western blot).
concentration was not affected by any compound indicating no
cytotoxicity as shown in Table 2. Next, we investigated NO inhibi-
tion by these compounds further to determine whether it was
caused by a reduced expression of iNOS. As shown in Figure 3,
levels of iNOS cells treated with compound 4 was dramatically
decreased which is consistent with the findings shown in NO pro-
duction (Fig. 2 and Table 1). This indicates that the reduced expres-
sion of iNOS due to these compounds exposure was responsible for
the inhibition of NO production.
IC50 values of compounds 1–8 were evaluated by using
GraphPad Prism 4.0 software and showed 10.79, 4.13, 13.27,
0.57, 11.84, 10.42, 1.90 and 0.99 lM, respectively (Table 2). From
the aforementioned pharmacological results, we can conclude that
2-aroylbenzofurans having 4-hydroxyphenyl moiety and/or
hydroxy (–OH) group at 5- and/or 6-position of benzofuran
scaffold are fruitful to show potent anti-inflammatory activity by
effective inhibition of iNOS with no cytotoxicity.
In summary, we have described an efficient synthesis of
rugchalcones A (1), B (2) and their derivatives (3–8) using substi-
tuted salicylaldehydes and phenacyl bromides as building blocks
and Rap–Stoermer reaction as a key step. Later, their anti-inflam-
matory effects were evaluated in lipopolysaccharide (LPS)
Table 1
anti-Inflammatory activities of rugchalcones A (1), B (2) and their derivatives (3–8)
Compound
NO production (% inhibition)
10
1
lM
lM
Medium (MED)
1.0 0.0 (99.0)⁄⁄⁄
83.8 6.0 (16.2)
100.0 2.7 (0.0)
92.2 0.5 (7.8)
45.8 1.8 (54.2)⁄⁄⁄
90.5 6.5 (9.5)
91.4 8.4 (8.6)
80.3 1.7 (19.7)
57.1 1.1 (42.9)⁄⁄⁄
79.1 4.1 (20.9)
1.0 0.0 (99.0)⁄⁄⁄
57.0 6.5 (43.0)⁄⁄⁄
0.7 0.7 (99.3)⁄⁄⁄
70.3 3.4 (29.7)⁄⁄
0.4 0.4 (99.6)⁄⁄⁄
70.2 4.7 (29.8)⁄⁄
64.2 3.5 (25.8)⁄⁄⁄
3.2 1.9 (96.8)⁄⁄⁄
25.7 0.2 (74.3)⁄⁄⁄
7.6 4.0 (92.4)⁄⁄⁄
1
2
3
4
5
6
7
8
L
-NMMA
The results are reported as mean value SEM for n = 3. Statistical significance is
based on the difference when compared with LPS-treated groups
(
**P <0.01,
***P <0.001).
% inhibition is based on LPS as shown in parenthesis.

1524
Y. H. Seo et al. / Bioorg. Med. Chem. Lett. 26 (2016) 1521–1524
3. (a) Duncan, A. J.; Heales, S. J. Mol. Aspects Med. 2005, 26, 67; (b) Bian, K.; Murad,
F. Front. Biosci. 2003, 8, d264.
stimulated RAW-264.7 macrophages. Of the 2-aroylbenzofurans
prepared in this study, four compounds, that is, compound 4
4. Hippisley-Cox, J.; Coupland, C.; Logan, R. Br. Med. J. 2005, 331, 1310.
5. (a) Hou, X.-L.; Yang, Z.; Wong, H. N. C. Prog. Heterocycl. Chem. 2002, 14, 139; (b)
Nevagi, R. J.; Dighe, S. N.; Dighe, S. N. Eur. J. Med. Chem. 2015, 97, 561.
6. Silva, D. H. S.; Pereira, F. C.; Zanoni, M. V. B.; Yoshida, M. Phytochemistry 2001,
57, 437.
(99.6%; IC50 = 0.57
7 (96.8%; IC50 = 1.90
exceptional inhibitory activity at 10
l
M), rugchalcone B (2) (99.3%; IC50 = 4.13
M) and 8 (74.3%; IC50 = 0.99 M) showed
M with no cytotoxicity. This
lM),
l
l
l
study revealed that compounds 2, 4, 7 and 8 which are having
4-hydroxyphenyl group and/or hydroxy (–OH) group at 5- and/or
6-position of benzofuran motif could be considered as a promising
scaffolds for the further development of NO inhibitors for potential
anti-inflammatory applications.
7. Sogabe, S.; Masubuchi, M.; Sakata, K.; Fukami, T. A.; Morikami, K.; Shiratori, Y.;
Ebiike, H.; Kawasaki, K.; Aoki, Y.; Shimma, N.; D’Arcy, A.; Winkler, F. K.; Banner,
D. W.; Ohtsuka, T. Chem. Biol. 2002, 9, 1119.
8. (a) Khan, W.; Alam, M. J.; Rashid, M.; Chowdhury, R. Bioorg. Med. Chem. 2005,
13, 4796; (b) Alper-Hayta, S.; Arisoy, M.; Temiz-Arpaci, O.; Yildiz, I.; Aki, E.;
Özkan, S.; Kaynak, F. Eur. J. Med. Chem. 2008, 43, 2568.
9. Day, S. H.; Chiu, N. Y.; Tsao, L. T.; Wang, J. P.; Lin, C. N. J. Nat. Prod. 2000, 63,
1560.
10. Filzen, G. F.; Bratton, L.; Cheng, X. M.; Erasga, N.; Geyer, A.; Lee, C.; Lu, G.;
Pulaski, J.; Sorenson, R. J.; Unangst, P. C.; Trivedi, B. K.; Xu, X. Bioorg. Med. Chem.
Lett. 2007, 13, 3630.
11. Manna, K.; Agrawal, Y. K. Eur. J. Med. Chem. 2010, 45, 3831.
12. (a) Rida, S. M.; El-Hawash, S. A. M.; Fahmy, H. T. Y.; Hazzaa, A. A.; El-Meligy, M.
M. M. Arch. Pharm. Res. 2006, 29, 826; (b) Hayakawa, I.; Shioya, R.; Agatsuma,
T.; Furukawa, H.; Naruto, S.; Sugano, Y. Bioorg. Med. Chem. Lett. 2004, 14, 4383.
13. Ono, M.; Saji, H. Med. Chem. Commun. 2015, 6, 391.
Acknowledgements
This research was financially supported by Priority Research
Centers Program through the National Research Foundation of
Korea (NRF) funded by the Ministry of Education, Science and
Technology (NRF-2009-0094071), South Korea.
14. (a) Oter, O.; Ertekin, K.; Kirilmis, C.; Koca, M.; Ahmedzade, M. Sens. Actuators, B:
Chem. 2007, 122, 450; (b) Qiu, J.-X.; Li, Y.-X.; Yang, X.-F.; Nie, Y.; Zhang, Z.-W.;
Chen, Z.-H.; Sun, G.-X. J. Mater. Chem. C 2014, 2, 5954.
Supplementary data
15. Du, G.; Han, J.-M.; Kong, W.-S.; Zhao, W.; Yang, H.-Y.; Yang, G.-Y.; Gao, X.-M.;
Hu, Q.-F. Bull. Korean Chem. Soc. 2013, 34, 1263.
16. (a) Kim, C. G.; Jun, J.-G. Bull. Korean Chem. Soc. 2015, 36, 2278; (b) Seo, Y. H.;
Kim, J.-K.; Jun, J.-G. Bioorg. Med. Chem. Lett. 2014, 24, 5727.
17. (a) Rap, E. Gazz. Chim. Ital. 1895, 25, 285; (b) Steormer, R. Liebigs Ann. Chem.
1900, 312, 237.
Supplementary data associated with this article can be found, in
the online version, at http://dx.doi.org/10.1016/j.bmcl.2016.02.
023.
References and notes
18. Kwon, K. H.; Murakami, A.; Hayashi, R.; Ohigashi, H. Biochem. Biophys. Res.
Commun. 2005, 337, 647.
19. (a) Chen, B.; Stout, R.; Campbell, W. F. FEMS Immunol. Med. Microbiol. 1996, 14,
109; (b) Salerno, L.; Sorrenti, V.; Di Giacomo, C.; Romeo, G.; Siracusa, M. A. Curr.
Pharm. Des. 2002, 8, 177; (c) Ueda, S.; Terauchi, H.; Yano, A.; Ido, M.;
Matsumoto, M.; Kawasaki, M. Bioorg. Med. Chem. Lett. 2004, 14, 313.
1. Quintans, J. Immunol. Cell Biol. 1994, 72, 262.
2. (a) Ruedi, K. B.; Christina, F.; Paul, N.; Michael, S.; Sterner-Kock, A.; Kock, M.;
Putney, L.; Ferrick, D. A.; Hyde, D. M.; Love, R. B. Inflammation 2008, 31, 167; (b)
Coussens, L. M.; Werb, Z. Nature 2002, 420, 860.