Antinociceptive properties of chalcones. Structure-activity relationships

Article abstract of DOI:10.1002/1521-4184(200110)334:10<332::AID-ARDP332>3.0.CO;2-O

Eleven chalcones were prepared and tested as antinociceptive agents using the writhing test in mice. Some compounds, given intraperitoneally, caused potent and dose-related antinociception, being several times more active than some reference drugs. The results evidenced that some physico-chemical parameters are involved in the pharmacological activity. 3,4-Dichlorochalcone (2) was the most effective compound, and was also studied in another model of pain in mice, the formalin test. Here it inhibited only the inflammatory pain (second phase), being equipotent to the reference drugs.

Full text of DOI:10.1002/1521-4184(200110)334:10<332::AID-ARDP332>3.0.CO;2-O

Notes
Rogério Corrêaa),
Antinociceptive properties of chalcones.
Structure-activity relationships
Márcia A. S. Pereiraa),
Daniela Buffona),
Lorena dos Santosa),
Valdir Cechinel Filhoa),
Adair R.S. Santosa),
Ricardo J. Nunesb)
Eleven chalcones were prepared and tested as antinociceptive agents using the writhing
test in mice. Some compounds, given intraperitoneally, caused potent and dose-related
antinociception, being several times more active than some reference drugs. The results
evidenced that some physico-chemical parameters are involved in the pharmacological
activity. 3,4-Dichlorochalcone (2) was the most effective compound, and was also studied
in another model of pain in mice, the formalin test. Here it inhibited only the inflammatory
pain (second phase), being equipotent to the reference drugs.
a)
Núcleo de Investigações Químico-
Farmacêuticas (NIQFAR)/CCS, Uni-
versidade do Vale do Itajaí (UNIVALI),
88302-202, Itajaí-SC, Brazil
Key Words: Chalcones; Antinociception; Writhing test; Mice
b)
Departamento de Química, Univer-
Received: May 15, 2001 [FP581]
sidade Federal de Santa Catarina
(UFSC), 88040-900, Florianópolis-SC,
Brazil
The results summarized in Table 1 indicate that several
chalcones exhibited potent antinociceptive effect when as-
Introduction
Chalcones or 1,3-diaryl-2-propen-1-ones are natural or
synthetic compounds belonging to the flavonoid family.
They exhibit different kinds of biological activities, such as
antimicrobial, anticancer, antiprotozoal, antiulcer, antiin-
flammatory, among others [1–4], and thus comprise a class
with important therapeutic potential. Previous studies per-
formed by Cechinel Filho and co-workers (1996) have
demonstrated that some simple chalcones derived from
xanthoxylin exert antinociceptive and antioedematogenic
action in mice, and were more potent than some well-known
antiinflammatory and analgesic drugs[5]. Such experimen-
tal observation and the fact that most of the actual analgesic
drugs cause undesired effects and are not useful in all
cases[6] encourage us to synthesize other chalcones with
possible antinociceptive properties. Thus, we have pre-
pared 11 compounds using a classical condensation reac-
tion (base-catalyzed) of substituted acetophenones and
benzaldehydes which were evaluated as antinociceptive
agents by using the writhing test in mice. The most active
compound was also analysed in the formalin-induced pain
test. Furthermore, we have preliminarily investigated the
structure-activity relationships of these compounds. The
results of some reference drugs were included for compari-
son.
sessed by the writhing test in mice. Chalcone4, which does
not contain substituent groups in either aromatic ring, gave
a calculated ID50 value of 99.7 µmol/kg, being equipotent
to aspirin and paracetamol, two standard drugs widely
employed in clinical practice. The introduction of different
substituent groups in the aromatic rings caused drastic
changes in activity. The choice of halogen atoms or methyl
group on the rings A and B of chalcones was made in order
to verify the influence of some physico-chemical parame-
ters, e.g. hydrophobicity, electronic, and steric effects.
In general, the introduction of substituent groups in the
ring A of chalcones produces more active compounds. The
two most potent compounds, chalcones 2 and 11 contain-
ing two or one chloro atom attached in the ring A, showed
Table 1. Effect of chalcones and some reference drugs given
intraperitoneally against acetic acid-induced abdominal constric-
tion (writhing test) in mice.
————————————————————————————–
Compound
ID50 (µmol/kg)
MIa (%)
————————————————————————————–
1
2
3
4
5
6
7
8
45.5 (23.3–81.5)b
9.0 (5.0–15.8)
90 ± 4.0
91 ± 4.0
92 ± 3.0
75 ± 5.0
65 ± 5.0
97 ± 2.0
54 ± 3.0
80 ± 1.0
95 ± 2.0
88 ± 3.0
93 ± 1.0
83 ± 2.0
88 ± 1.0
93 ± 7.0
16.8 (7.3–38.4)
99.7 (44.2–224.6)
108.3 (64.9–176.9)
16.0 (11.8–21.8)
35.5 (28.8–43.6)
113.4 (76.7–167.8)
113.9 (72.3–180.6)
137.8 (103.1–184.2)
13.0 (8.8–19.5)
Results and discussion
The scheme shows the general synthetic procedure used.
All the compounds were obtained in good yields (70–98%)
and readily characterized by conventional spectral data.
Inspection of the 1H NMR spectra suggested that the
chalcones were geometrically pure and configured trans
(JHα-Hβ = 15–16 Hz)[4].
9
10
11
Aspirinc
Paracetamolc
Diclofenacd
133.2 (73.0–243.1)
125.0 (104.0–150.0)
38.0 (29.5–49.0)
———
———————————————–—————————————
a maximal inhibition; b 95% confidence limit; c from reference [7];
d from reference [12]. Each group represents the mean of five to
seven animals.
Correspondence: Prof. Valdir Cechinel Filho, Núcleo de Investi-
gações Químico-Farmacêuticas (NIQFAR)/CCS, Universidade do
Vale do Itajaí (UNIVALI), 88302-202, Itajaí-SC, Brazil.
E-mail: cechinel@univali.br
© WILEY-VCH Verlag GmbH, D-69451 Weinheim, 2001
0365-6233/01/1010/0332 $17.50 +.50/0

Arch. Pharm. Pharm. Med. Chem. 2001, 334, 332–334
Antinociceptive properties of chalcones 333
Experimental
Y
O
C
O
C
O
C
H
CH3
+
B
CH CH
1) NaOH/EtOH
2) AcOH/H
A
Synthesis: general procedures
O
2
X
Y
X
Compounds 1–11 were obtained by reaction of acetophenone
and benzaldehyde (1:1) in the presence of sodium hydroxide
and ethanol and further addition to cool diluted acetic acid,
according the methodology previously described[8]. The re-
spective products were purified by recrystallization or column
chromatography over silica gel. All the compounds were syn-
thesized in good yields (55–98%) and characterized by
1H NMR, IR and microanalyses. The purity of these compounds
was determined by TLC using several solvent systems of dif-
ferent polarity.
X=4-Br; Y=H (1)
X= 3,4-Cl ; Y= H (2)
2
X= 3,4-Cl ; Y= 4-N(CH
) (3)
3 2
2
X= Y= H (4)
X= Y= 4-Cl (5)
X= 3,4-Cl ; Y= 4-Cl (6)
2
X= 4-CH ; Y= 4-Cl (7)
3
X= H; Y= 4-Cl (8)
X= 4-Br; Y= 4-Cl (9)
X= 4-CH ; Y= H (10)
3
X= 4-Cl; Y=H (11)
Scheme
ID50 values of 9.0 and 13.0 µmol/kg, and maximal inhibition
of 91 and 93%, respectively. They were about 14-fold more
active than aspirin and paracetamol and about 4-fold more
potent than diclofenac in the writhing test. It should be noted
that the introduction of a methyl group (chalcone 10) in-
stead of halogen atoms in the ring A considerably de-
creased the antinociceptive potency, suggesting that
electron-withdrawing groups but not electron-donor groups
increase the antinociceptive effect. This hypothesis may be
supported by the activity of compound 1, which was about
3-fold more active than compound 10. However, other
additional studies are required to confirm such observation.
Although both bromine or chloro atoms present in the
position 4 of ring A improve the pharmacological effect, the
latter seems to be more effective, indicating that a steric
parameter is also involved in the antinociceptive activity of
these compounds. In addition, preliminary evaluation of the
hydrophobicity by using the periodic box method by Hyper
Chem indicated that this parameter is not related with the
antinociceptive activity of these compounds.
Physico-chemical data of synthesized compounds
Compound (1): 1H-NMR (DMSO-d6, ppm): 7.27 (d, 1H,
CH=CH), 7.38 (d, 1H, CH=CH, J = 15.63), 7.51–7.75 (m, 9H,
Ar); FT-IR (KBr disk, cm–1): 1660 (ν -C=O), 1605 (ν -C=C-); mp
(°C): 155–160; Yield: 54.8%.
Compound (2): 1H-NMR (DMSO-d6 , ppm): 7.27 (d, 1H,
CH=CH), 7.39 (d, 1H, CH=CH, J = 15.70), 7.50–7.93 (m, 8H,
Ar); FT-IR (KBr disk, cm–1): 1661 (ν -C=O), 1605 (ν -C=C-); mp
(°C): 96–100; Yield: 68%.
Compound (3): 1H-NMR (DMSO-d6, ppm): 3.01 (s, 6H, N-
(CH3)2), 7.28 (d, 1H, CH=CH), 7.41 (d, 1H, CH=CH, J = 15.71),
7.51–7.75 (m, 7H, Ar); FT-IR (KBr disk, cm–1): 1649 (ν -C=O),
1612 (ν -C=C-); mp (°C): 124–126; Yield: 85%.
Compound (4): 1H-NMR (DMSO-d6, ppm): 6.38 (d, 1H,
CH=CH), 6.89 (d, 1H, CH=CH, J = 15.72), 7.25–7.76 (m, 10 H,
Ar); FT-IR (KBr disk, cm–1): 1661 (ν -C=O), 1605 (ν -C=C-); mp
(°C): 119–122; Yield: 77%.
Compound (5):1H-NMR (DMSO-d6, ppm): 7.28 (d, 1H,
CH=CH), 7.42 (d, 1H, CH=CH, J = 15.71), 7.47–7.87 (m, 8H,
Ar); FT-IR (KBr disk, cm–1): 1657 (ν -C=O), 1605 (ν -C=C-), mp
(°C): 160–163; Yield: 89%.
In view of the interesting antinociceptive activity of chalcone
2, it was analysed by the formalin-induced pain test, which
defines two distinct periods of response, i.e., “early re-
sponse (neurogenic pain)” and “late response (inflamma-
tory pain)”. It dose-dependently prevented only the late
phase of the formalin test, like the standard drugs. The
calculated ID50 value was 111.5 (50.5–243.7) µmol/kg with
maximal inhibition of 82%, being equipotent to aspirin and
acetaminophen, which presented ID50 of approximately
120 µmol/kg in relation to the second phase of the formalin
Compound (6): 1H-NMR (DMSO-d6, ppm): 7.29 (d, 1H,
CH=CH), 7.43 (d, 1H, CH=CH, J = 15.69), 7.54–7.94 (m, 7H,
Ar); FT-IR (KBr disk, cm–1): 1661 (ν -C=O), 1602 (ν -C=C-); mp
(°C): 120–123; Yield: 96%.
Compound (7): 1H-NMR (DMSO-d6, ppm): 2.43 (s, 3H, CH3),
7.51 (d, 1H, CH=CH), 7.41 (d, 1H, CH=CH, J = 15.70), 7.44–
7.80 (m, 8H, Ar); FT-IR (KBr disk, cm–1): 1657 (ν -C=O), 1605
(ν -C=C-); mp (°C): 159–162; Yield: 80%.
test [7]
.
Compound (8): 1H-NMR (DMSO-d6, ppm): 7.28 (d, 1H,
CH=CH), 7.42 (d, 1H, CH=CH, J = 15.71), 7.25–7.76 (m, 9H,
Ar); FT-IR (KBr disk, cm–1): 1658 (ν -C=O), 1604 (ν -C=C-); mp
(°C): 122–125; Yield: 74%.
The mechanism by which the chalcones exert antinocicep-
tive action still remains undetermined, but chemical and
pharmacological studies are currently in progress in our
laboratories and the results will be published elsewhere.
Compound (9): 1H-NMR (DMSO-d6, ppm): 7.28 (d, 1H,
CH=CH), 7.43 (d, 1H, CH=CH, J = 15.69), 7.54–7.75 (m, 8H,
Ar); FT-IR (KBr disk, cm–1): 1657 (ν -C=O), 1604 (ν -C=C-); mp
(°C): 168–170; Yield: 97%.
Considering that only a few authors have demonstrated the
antinociceptive activity of chalcones and the level of current
interest in discovering other medicinal agents to treat pain
processes, we believe that some compounds, such as
chalcones 2 and 11, might be further used as models to
obtain new more potent and/or selective analgesic drugs.
Compound (10): 1H-NMR (DMSO-d6,ppm): 2.42 (s, 3H, CH3),
7.93 (d, 1H, CH=CH), 7.37 (d, 1H, CH=CH, J = 15.65), 7.43–
7.79 (m, 9H, Ar); FT-IR (KBr disk, cm–1): 1656 (ν -C=O), 1596
(ν -C=C-); mp (°C): 95.6–100; Yield: 96%.
Compound (11): 1H-NMR (DMSO-d6, ppm): 7.28 (d, 1H, CH=CH);
7.39 (d, 1H, CH=CH, J = 15.72), 7.47–7.78 (m, 9H, Ar); FT-IR (KBr
disk, cm–1): 1661 (ν -C=O), 1606 (ν -C=C-); mp (°C): 85–92; Yield:
98%.
Acknowledgements
This work was supported by grants from CNPq and ProP-
PEx/UNIVALI.

334 Antinociceptive properties of chalcones
Arch. Pharm. Pharm. Med. Chem. 2001, 334, 332–334
Pharmacological analysis: evaluation of antinociceptive
activity
Statistical analysis
The results are presented as mean ± s.e.m., and the statistical
significance between the groups was analysed by means of an
analysis of variance followed by Dunnett’s multiple comparison
test. P values less than 0.05 were considered as indicative of
significance. The ID50 values (the dose of the compound that
reduced responses by 50% in relation to control values) were
estimated by graphical interpolation from individual experi-
ments. ID50’s are presented as mean values and 95% confi-
dence interval. MI is the maximum inhibition at higher dose
used.
Writhing test
Male Swiss mice (25–30 g) were used. The abdominal constric-
tion induced by intraperitoneal injection of acetic acid (0.6%),
was carried out according to the procedures described pre-
viously[9,10] with minor modifications. The animals were pre-
treated with chalcones intraperitoneally 30 min before the acid
acetic injection. Control animals received a similar volume of
0.9% NaCl (10 ml kg–1 , i.p.). All experiments were carried out
at 23 ± 2 °C. The pairs of mice were placed in separate boxes
and the number of abdominal constrictions of the abdominal
muscles together with a stretching, were cumulatively counted
over a period of 20 min. Antinociceptive activity was expressed
as the reduction of the number of abdominal contractions
between control animals and mice pretreated with the com-
pound studied.
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