Synthesis, characterization, and cytotoxic activities of heterocyclic chalcones containing furan, and crystal structure of 1-(4-iodophenyl)-3-(5-methylfuran-2-yl)prop-2-en-1-one

Article abstract of DOI:10.1080/15421406.2016.1149025

A series of heterocyclic chalcone (3a–e, 5, 7) were synthesized and characterized by Infrared, ¹H and ¹³C nuclear magnetic resonance, and mass spectra. Crystal structure of 1-(4-iodophenyl)-3-(5-methylfuran-2-yl)prop-2-en-1-one was d

Full text of DOI:10.1080/15421406.2016.1149025

Molecular Crystals and Liquid Crystals
ISSN: 1542-1406 (Print) 1563-5287 (Online) Journal homepage: http://www.tandfonline.com/loi/gmcl20
Synthesis, characterization, and cytotoxic activities
of heterocyclic chalcones containing furan,
and crystal structure of 1-(4-iodophenyl)-3-(5-
methylfuran-2-yl)prop-2-en-1-one
Zakiah Mokhtar, Joazaizulfazli Jamalis, Siti Pauliena Mohd Bohari, Mohd
Mustaqim Rosli & Hoong-Kun Fun
To cite this article: Zakiah Mokhtar, Joazaizulfazli Jamalis, Siti Pauliena Mohd Bohari, Mohd
Mustaqim Rosli & Hoong-Kun Fun (2016) Synthesis, characterization, and cytotoxic activities
of heterocyclic chalcones containing furan, and crystal structure of 1-(4-iodophenyl)-3-(5-
methylfuran-2-yl)prop-2-en-1-one, Molecular Crystals and Liquid Crystals, 631:1, 119-131, DOI:
10.1080/15421406.2016.1149025
To link to this article: http://dx.doi.org/10.1080/15421406.2016.1149025
Published online: 12 Jul 2016.
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Date: 17 July 2016, At: 22:40

MOLECULAR CRYSTALS AND LIQUID CRYSTALS
, VOL. , –
http://dx.doi.org/./..
Synthesis, characterization, and cytotoxic activities of
heterocyclic chalcones containing furan, and crystal structure of
-(-iodophenyl)--(-methylfuran--yl)prop--en--one
Zakiah Mokhtara, Joazaizulfazli Jamalisa, Siti Pauliena Mohd Boharib, Mohd
Mustaqim Roslic, and Hoong-Kun Func,d
aDepartment of Chemistry, Faculty of Science, Universiti Teknologi Malaysia, UTM Johor Bahru, Johor, Malaysia;
bDepartment of Biotechnology and Medicinal Engineering, Faculty of Bioscience and Medical Engineering,
Universiti Teknologi Malaysia, UTM Johor Bahru, Johor, Malaysia; cX-ray Crystallography Unit, School of Physics,
Universiti Sains Malaysia, Penang, Malaysia; dDepartment of Pharmaceutical Chemistry, College of Pharmacy,
King Saud University, Riyadh, Saudi Arabia
ABSTRACT
KEYWORDS
Synthesis; breast cancer;
crystal structure; cytotoxicity;
heterocyclic chalcone
A series of heterocyclic chalcone (3a–e, 5, 7) were synthesized and
characterized by Infrared, 1H and 13C nuclear magnetic resonance, and
mass spectra. Crystal structure of 1-(4-iodophenyl)-3-(5-methylfuran-2-
yl)prop-2-en-1-one was determined using single crystal X-ray diffrac-
tion. All the synthesized compounds were evaluated for their cytotoxic
activities on MDA MB 231 and CHO cell by using MTT assay. Compound
3c showed good cytotoxic effect on MDA MB 231 with IC50 values of
9.8 µg/mL.
Introduction
Cancer is the second cause of human death in the world after cardiovascular diseases [1]. It
reoccurs due to the presence of malignant tumor in the body with an ability to invade and
spread to other tissues [2]. Breast cancer, for example, is one of the most common cancers
among women [3, 4] and the first of the most frequent in Malaysia followed by colorectal and
lung cancers [5]. There are various types of treatments, such as chemotherapy, radiotherapy,
hormone therapy, surgery [6], and hyperthermia [7], offered to patients diagnosed with can-
cer. However, some of the treatments resulted with undesirable side effects to patients such as
nausea, vomiting, poor oral intake, cachexia, and lethargy [8].
Bacterial infections are other types of health problems infect humans around the world [9].
The emergence of antibiotics to fight the microbes and other bacteria has resulted in the pres-
ence of pathogens that are resistant to antibiotics. The presence of drugs, such as amoxicillin,
norfloxacin, and ciprofloxacin, can help to fight against the bacterial infection. Nevertheless,
these drugs have side effects such as dizziness, nausea, hypertension, et al. [9, 20].
CONTACT Joazaizulfazli Jamalis
joazaizul@kimia.fs.utm.my
Teknologi Malaysia,  UTM Johor Bahru, Johor, Malaysia.
Department of Chemistry, Faculty of Science, Universiti
Color versions of one or more of the figures in the article can be found online at www.tandfonline.com/gmcl.
Crystallographic data for compound 3e reported in this paper have been deposited at the Cambridge Crystallographic Data
Centre (CCDC) with CCDC number .
©  Taylor & Francis Group, LLC

120
Z. MOKHTAR ET AL.
Therefore, chalcone can be one of the compounds that can act in tumor cell and fight
against bacteria as it exhibit a broad range of pharmacological activities such as antibacterial,
anticancer, antifungal, anti-inflammatory, antioxidant, et al. [10]. Secondary metabolites [11]
are found abundantly in nature from fern to higher plants. The presence of α,β-unsaturated
carbonyl group in its compound is the indication of the formation of chalcone from two
aromatic rings, and the removal of this group could obstruct its various pharmacological
activities [8].
Based on its diversity of biological activities, chalcone has attracted our attention to synthe-
size instead of isolating it from natural sources, as the extraction method is time-consuming
and only small amount of product is formed. Previous study carried out by Solomon and Lee
[4] reported that heteroarylchalcone showed an improved anti-proliferative activity on cancer
cells than the commonly prescribed cisplatin. Encouraged by this, a series of heterocyclic chal-
coneanalogues were synthesized by focusing on the heterocyclic electron-rich oxygen with
different substituents of phenyl ring. All the compounds were tested for their ability to act as
antibacterial and anticancer agents to develop a novel therapeutic agent for better treatment
of cancer and fight bacterial infections.
Materials and methods
5-Methylfurfural (99%), 4-chloroacetophenone (97%), 4-fluoroacetophenone (99%), 2-
acetylpyrazine (ࣙ99%), 4-acetylpyridine (97%), MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-
diphenyltetrazolium bromide] reagent, and pyridine (>99%, Fluka) were purchased from
Sigma, Subang Jaya, Selangor, Malaysia. Chemical reagents, 4-bromoacetophenone (98%,
Acros Organic) and 4-iodoacetophenone (98%, Acros Organic) were purchased from Fischer
Scientific, Shah Alam, Selangor, Malaysia. Benzaldehyde (>99%) and dimethlyamine were
bought from Merck, Malaysia. Sodium hydroxide pellets, potassium hydroxide pellets, and
hydrochloric acid (37%) were of analytical grade and obtained from Qrec, Rawang, Selangor,
Malaysia. Melting points (uncorrected) were determined using a Barnstead Electrothermal
9100 melting point apparatus. Infrared (IR) spectra were recorded on a Perkin-Elmer Uni-
versal ATR spectrophotometer. 1H and 13C nuclear magnetic resonance (NMR) spectra were
performed on BrukerAvance spectrometer at 400 MHz. The chemical shifts were recorded
in parts per million (δ) in deuterochloroform solution. Results of mass spectra (MS) were
obtained from NUS Mass Spectrometry Service, Singapore. Reactions were monitored using
precoated silica gel 60 F254 TLC plates and observed under ultraviolet (UV)254 light.
Synthesis of chalcones (3a–e)
A mixture of 5-methyl-2-furaldehyde (21.72 mmol) with substituted acetophenone (21.72
mmol) was dissolved in absolute ethanol (20 mL). Slowly, aqueous NaOH solution (4.3 mL,
2%) was added drop-wise [12] at 0°C and stirred for 24 hr at 25°C. After the completion of
reaction was tested on pre-coated TLC plates, ice water (100 mL) was added and neutralized
with 10% HCl. The precipitate formed was collected by vacuum filtration, washed with cold
water, dried, and recrystallized using ethanol.
-(-Methylfuran--yl)--phenylprop--en--one (3a) []
Bright yellow powder (70.1% yield), m.p. 64.1–66.0°C, IR (cm−1): 3125 (C-H aromatic
stretching), 2920 (C-H aliphatic), 1653 (C=O), 1586 (C=C olefinic), 1551 and 1444 (C=C
1
aromatic). H NMR (400 MHz, CDCl3): δ 2.42 (3H, s, CH3-6ꢀ), 6.16 (1H, dd, J = 0.8 and
3.2 Hz, H-4ꢀ), 6.65 (1H, d, J = 3.2 Hz, H-3ꢀ), 7.39 (1H, d, J = 15.2 Hz, H-2), 7.50 (3H, m,
phenyl-H), 7.55 (1H, d, J = 15.2 Hz, H-3), 8.05 (2H, m, phenyl-H).

MOLECULAR CRYSTALS AND LIQUID CRYSTALS
121
-(-Fluorophenyl)--(-methylfuran--yl)prop--en--one (3b) []
Yellow needles (88.6% yield), m.p. 65.6–68.9°C, IR (cm−1): 3097 (C-H aromatic), 2924 (C-
H aliphatic), 1657 (C=O), 1601 (C=C olefinic), 1563 and 1504 (C=C aromatic). 1H NMR
(400 MHz, CDCl3): δ 2.42 (3H, s, CH3-6ꢀ), 6.16 (1H, dd, J = 0.8, and 3.2 Hz, H-4ꢀ), 6.66 (1H,
d, J = 3.2 Hz, H-3ꢀ), 7.16 (2H, m, phenyl-H), 7.35 (1H, d, J = 15.2 Hz, H-2), 7.55 (1H, d, J =
15.2 Hz, H-3), 8.07 (2H, m, phenyl-H).
-(-Chlorophenyl)--(-methylfuran--yl)prop--en--one (3c) []
Yellow crystals (85.3% yield), m.p. 104.7–107.8°C, IR (cm−1): 3113 (C-H aromatic), 2921 (C-
H aliphatic), 1655 (C=O), 1597 (C=C olefinic), 1562 (C=C aromatic). 1H NMR (400 MHz,
CDCl3): δ 2.42 (3H, s, CH3-6ꢀ), 6.16 (1H, dd, J = 0.8 and 3.2 Hz, H-4ꢀ), 6.67 (1H, d, J = 3.2 Hz,
H-3ꢀ), 7.33 (1H, d, J = 15.2 Hz, H-2), 7.48 (2H, br d, J = 8.4 Hz, H-3ꢀꢀ, H-5ꢀꢀ), 7.55 (1H, d, J =
15.2 Hz, H-3), 7.99 (2H, br d, J = 8.4 Hz, H-2ꢀꢀ, H-6ꢀꢀ).
-(-Bromophenyl)--(-methylfuran--yl)prop--en--one (3d) []
Yellow crystal (90% yield), m.p. 109.6–111.9°C, IR (cm−1): 3110 (C-H aromatic), 2921 (C-
H aliphatic), 1654 (C=O), 1598 (C=C olefinic), 1560 (C=C aromatic). 1H NMR (400 MHz,
CDCl3): δ 2.42 (3H, s, CH3-6ꢀ), 6.16 (1H, dd, J = 0.8 and 3.2 Hz, H-4ꢀ), 6.67 (1H, d, J = 3.2 Hz,
H-3ꢀ), 7.32 (1H, d, J = 15.2 Hz, H-2), 7.55 (1H, d, J = 15.2 Hz, H-3), 7.64 (2H, br d, J = 8.4 Hz,
H-3ꢀꢀ, H-5ꢀꢀ), 7.91 (2H, br d, J = 8.4 Hz, H-2ꢀꢀ, H-6ꢀꢀ).
-(-iodophenyl)--(-methylfuran--yl)prop--en--one (3e)
Orange crystal (71.7% yield), m.p. 101.8–103.0°C, IR (cm−1): 3108 (C-H aromatic), 2917 (C-
H aliphatic), 1651 (C=O), 1599 (C=C olefinic), 1559 (C=C aromatic). 1H NMR (400 MHz,
CDCl3): δ 2.42 (3H, s, CH3-6ꢀ), 6.16 (1H, dd, J = 0.8 and 3.2 Hz, H-4ꢀ), 6.67 (1H, d, J = 3.2 Hz,
H-3ꢀ), 7.32 (1H, d, J = 15.2 Hz, H-2), 7.55 (1H, d, J = 15.2 Hz, H-3), 7.75 (2H, br d, J = 8.4 Hz,
H-3ꢀꢀ, H-5ꢀꢀ), and 7.86 (2H, br d, J = 8.4 Hz, H-2ꢀꢀ, H-6ꢀꢀ). 13C NMR (100 MHz, CDCl3): δ
14.04 (C-6ꢀ), 100.32 (C-1ꢀꢀ), 109.52 (C-4ꢀ), 116.82 (C-3), 118.76 (C-3ꢀ), 129.83 (C-3ꢀꢀ, C-5ꢀꢀ),
131.25 (C-2), 137.84 (C-2ꢀꢀ, C-6ꢀꢀ), 150.26 (C-2ꢀ), 156.22 (C-5ꢀ), 188.98 (C=O). Found (ESI):
338.9876 [M+ H+], (C14H12IO2 requires 338.9882).
Synthesis of -(-methylfuran--yl)--(pyrazin--yl)prop--en--one (5) []
To a solution of 5-methyl-2-furaldehyde (24.59 mmol) and 2-acetylpyrazine (24.59 mmol)
was added 20 mL of ethanol. The mixture was dissolved first before addition of 2% ethanolic
KOH (9.63 mL) at 0°C and stirred for 24 hr at 25°C. After the completion of reaction was
monitored by TLC, crushed ice was added to the reaction mixture. The precipitate formed
was filtered, washed with cold water, dried, and recrystallized by absolute ethanol.
-(-methylfuran--yl)--(pyrazin--yl)prop--en--one (5)
Yellow crystal (77.4% yield) m.p. 113.4–115.7°C, IR (cm−1): 3104 (C-H aromatic), 2923 (C-
H aliphatic), 1662 (C=O), 1598 (C=C olefinic), 1558 and 1518 (C=C aromatic). 1H NMR
(400 MHz, CDCl3): Table 1); 13C NMR (100 MHz, CDCl3): Table 1; Found (ESI): 215.0815
[M+ H+], (C12H11N2O2 requires 215.0821).
Synthesis of -(-methylfuran--yl)--(pyridine--yl)prop--en--one (7) []
5-Methyl-2-furaldehyde (8.26 mmol) was dissolved in 2 mL of pyridine before addition of
4-acetylpyridine (4.13 mmol) and dimethylamine (8.26 mmol). The reaction mixture was
stirred at room temperature for 24 hr. After the completion of reaction was tested by TLC,

122
Z. MOKHTAR ET AL.
it was poured into 100 mL of water and kept at 4°C overnight. The solid formed was filtered,
washed with cold water, dried, and recrystallized using ethanol.
-(-Methylfuran--yl)--(pyridine--yl)prop--en--one (7)
Black solid (73.9% yield), m.p. 103.5–107.3°C, IR (cm−1): 3116 (C-H aromatic), 2914 (C-
H aliphatic), 1661 (C=O), 1597 (C=C olefinic), 1551 (C=C aromatic). 1H NMR (400 MHz,
CDCl3): δ 2.44 (3H, s, CH3-6ꢀ), 6.20 (1H, dd, J = 0.4 and 3.2 Hz, H-4ꢀ), 6.75 (1H, d, J =
3.2 Hz, H-3ꢀ), 7.28 (1H, d, J = 15.2 Hz, H-2), 7.58 (1H, d, J = 15.2 Hz, H-3), 7.88 (2H, d, J
= 6.4 Hz, H-2ꢀꢀ, H-6ꢀꢀ), 8.85 (2H, d, J = 6.4 Hz, H-3ꢀꢀ, H-5ꢀꢀ). 13C NMR (100 MHz, CDCl3):
δ 14.05 (C-6ꢀ), 109.79 (C-4ꢀ), 116.48 (C-3), 119.71 (C-3ꢀ), 121.43 (C-2ꢀꢀ, C-6ꢀꢀ), 132.25 (C-2),
144.65 (C-2ꢀꢀ), 150.01 (C-3ꢀꢀ, C-5ꢀꢀ), 150.72 (C-2ꢀ), 156.85 (C-5ꢀ), 189.01 (C=O). Found (ESI):
214.0863 [M+H+], (C13H12NO2 requires 214.0868).
Cell lines
The human breast cancer cell (MDA MB 231) and Chinese Hamster Ovary (CHO) cell were
obtained from Tissue Engineering Laboratory, Faculty of Bioscience and Medical Engineer-
ing, UTM as a gift from Dr. Salehhuddin Hamdan. MDA MB 231 cell was cultured in Dul-
becco’s Modified Eagle Medium (DMEM), and CHO cell was cultured in Roswell Park Media
Institute (RPMI 1640). Both media were supplemented with 10% fetal bovine serum (FBS),
100 IU/mL of penicillin, and 100 µg/mL of streptomycin (GIBCO, Bio-Diagnostics, Petaling
Jaya, Selangor, Malaysia). The cells were maintained at 37°C in 5% CO2 (Panasonic, model
MCO-175 series).
MTT assay
The viability of cell was carried out using MTT assay. MDA MB 231 and CHO cells were
cultured in 96-well plates with a seeding density of 5 × 104 cells/mL, and were allowed to stay
for 24 hr in CO2 incubator. All chalcones were dissolved in dimethyl sulfoxide (DMSO) and
diluted with different concentrations ranging from 0.39 to 50 µg/mL. The final percentage
of DMSO used did not exceed 0.1%. Each sample, 100 µL, was added into 96-well plate, and
incubated for 72 hr. The cells were then treated with 20 µL of MTT reagent (5 mg/1 mL in
phosphate buffer solution (PBS)) into each well and incubated for an additional 4 hr. Buffer
solution, 225 µL (10% 0.1 M HCl and 90% isopropanol), was added in each well to solubilize
the formation of purple formazan produce by viable cells. The amount of formazan produced
was measured at 575 nm and 570 nm (as reference) recorded by microplate reader (BioTek
Instruments Inc., model Epoch, USA) [5, 16].
X-ray crystallography
X-ray crystallography analysis was performed to provide important information for future
structure–activity relationship studies as a compliment to NMR results. However, from the
seven studied chalcone derivatives, we were able to obtain a good crystal quality suitable for
X-ray crystallography analysis for compound 3e only. X-ray diffraction data for compound 3e
were collected from single crystal by using a Bruker APEX DUO CCD diffractometer with a
graphite monochromatic Mo-Kα radiation at a detector distance of 5 cm with APEX2 software
[17]. The collected data were reduced by using SAINT program and the empirical absorption
corrections were performed with the SADABS program [17]. The structure of compound 3e

MOLECULAR CRYSTALS AND LIQUID CRYSTALS
123
O
N
O
H3C
H
O
2a; R= H
2b; R= F
2c; R= Cl
2d; R= Br
2e; R = I
(1)
(6)
CH3
O
CH3
O
Pyridine,
Dimethylamine
R
aq. NaOH,
N
(2a-e)
CH3
Ethanol
O
N
(4)
O
CH3
O
ethanolic KOH,
Ethanol
O
CH3
N
R
(7)
(3a-e)
O
O
N
N
CH3
3a; R= H
3b; R= F
3c; R= Cl
3d; R= Br
3e; R = I
(5)
Scheme . Route of synthesis for the preparation of chalcone derivatives.
was solved by direct methods and was refined using a full-matrix least-squares method on
F2 using the SHELXTL program [18]. All non-hydrogen atoms were refined anisotropically.
˚
All hydrogen atoms were placed in calculated positions with C–H = 0.95–0.98 A and refined
using a riding model with Uiso(H) = 1.2 Ueq(C) and Uiso(H) = 1.5 Ueq(C-methyl). A rotating
group model was used for methyl groups. The final refinement converged well. Materials for
publication were prepared by using SHELXTL [18] and PLATON [19].
Results and discussion
Chemistry
In the current study, seven chalcone derivatives (3a–3e, 5, and 7) were successfully syn-
thesized in high yield under the Claisen–Schmidt’s condensation by reaction between
5-methyl-2-furaldehyde and different types of substituents of aromatic ring using basic con-
ditions as depicted in Scheme 1. The confirmations of the structure of compounds were
based on their spectral and physical data. Compound 5 is a new compound, synthesized from

Table . H and C NMR data for compound 5.
Position
δ (H)
δ (C)
COSY
HMBC



.
.
.
.
.
.
.
.
.
.
.
.
. (d, J = . Hz)
H-
H-
C-ꢀ, C-, C-
C-ꢀ, C-
. (d, J = . Hz)
ꢀ
ꢀ
ꢀ
ꢀ
ꢀ
ꢀꢀ
ꢀꢀ
ꢀꢀ
ꢀꢀ
-
. (d, J = . Hz)
H-ꢀ
H-ꢀ
C-ꢀ, C-ꢀ, C-ꢀ
C-ꢀ, C-ꢀ, C-ꢀ
. (dd, J = . and . Hz)
-
.
C-ꢀ, C-ꢀ
. (d, J = . Hz)
. (dd, J = . and . Hz)
. (d, J = . Hz)
C-ꢀ
C-ꢀ
C-ꢀ
δ: ppm

124
Z. MOKHTAR ET AL.
O
3
2' O
6'
6''
5''
N
N
5' CH3
2
3'
3''
4'
Figure . HMBC experiment for compound 5.
2-acetylpyrazine and 5-methyl-2-furaldehyde under basic conditions. This compound was
characterized by IR, 1H NMR, 13C NMR, 2D NMR, and MS.
The IR spectrum for compound 5 showed two stretching bands at 3104 and 2923 cm−1,
which were attributed for C-H sp2 aromatic and sp3 of methyl group respectively. The absorp-
tion band at 1662 cm−1 was assigned for carbonyl group, which indicated the formation of
chalcone. This band was slightly shifted to a lower frequency due to the conjugation effect
with the presence of aryl group and also α,β double bond on enone linkage at 1598 cm−1 [9].
The stretching frequency for C=C aromatic was detected at 1558 and 1518 cm−1, while for
C-H vinyl, it was observed at 1015 cm−1. These absorption bands were present in compounds
3a–e and 7 as they all have the same skeleton.
The condensation reaction for the formation of chalcone was proved successful by 1H NMR
spectrum. Compound 5 exhibited two doublet signals at δ 7.68 (d, J = 15.6 Hz) and δ 7.93
(d, J = 15.6 Hz), which were assignable for H-2 and H-3 respectively. The presence of these
two signals gave evidence of the presence of ethylene moiety in the enone linkage in trans-
conformation. The signal for the carbonyl group could be found farther downfield at δ 188.27.
These signals can also be found in other synthesized compounds (3a–e and 7).
The data from 1H NMR were supported with 13C NMR as it showed the presence of 12 car-
bon atoms (Table 1), which were represented by one methyl, three quaternary carbon atoms,
four methane carbons, three unequivalent carbon atoms of aromatic rings, and one carbonyl
carbon. The heteronuclear multiple quantum correlation (HMQC) data showed correlation
between proton and its carbon atoms. In this experiment, ethylene group, as a significant
indication of chalcone formation, showed correlation between 7.68 and 131.54, and 7.93 and
116.04, while the connection observed between 6.18 and 109.68, and 6.75 and 119.40 were
assigned for two methane carbons in the furan ring. The confirmation for the position such as
quaternary carbon atoms could be observed through the heteronuclear multiple bond correla-
tion (HMBC) experiment (Figure 1), which showed correlation of connected bonds between
proton and carbon separated by two or three bonds. The results from MS (ESI) gave evidence
of the formation of targeted heterocyclic compound with m/z = 215.0815, matched with its
molecular formula C12H10O2N2.
Table . Cytotoxic activity of heterocyclic chalcone derivatives on cancer and non-cancer cells.
Compoundsa
R
IC (µg/mL)b
CHO
.
.
.
.
.
.
.
MDA MB 
.
3a
3b
3c
3d
3e
5
Phenyl
-FPh
-ClPh
-BrPh
-IPh
-pyrazyl
-pyridyl
.
.
.
.
.
.
7
a
The structure of these compounds can be referred in Scheme .
IC: growth inhibition.
b

MOLECULAR CRYSTALS AND LIQUID CRYSTALS
125
Figure . Morphologic changes in CHO cell on treatment with heterocyclic chalcone: (a) cell without treat-
ment; (b) 3a; (c) 3b; (d) 3c; (e) 3d; (f) 3e; (g) 5; (h) 7, based on their IC values at  hr. The red arrow in the
picture of cells showed the morphology of cell.
Heterocyclic chalcone 7 was synthesized first by using the same basic condition as used for
compounds 3a–e. However, this condition has resulted in the failure of chalcone formation.
It was in agreement with Szües et. al. [14], who synthesized this chalcone using other acid
and basic condition but still had no improvement, and therefore they replaced the reagent
with diethylamine, which acted as a catalyst. The basicity of diethylamine is sufficient to form
the desired chalcone and avoid the formation of a complex product. However, in this study,
we use dimethylamine instead of diethylamine. Although its basicity is slightly lower than
diethylamine, changing the catalyst could still give the desired chalcone.
Cytotoxicity study of heterocyclic chalcones
Each synthesized compound was tested for its toxicity effects against breast cancer cell line
(MDA MB 231) and Chinese hamster ovary (CHO) cell as a positive control cell group. The
toxicity of the investigated compounds was expressed in IC50 values as summarized in Table 2.
Figure . Morphologic changes in MDA MB  cell on treatment with heterocyclic chalcone: (a) cell without
treatment; (b) 3a; (c) 3b; (d) 3c; (e) 3d; (f) 3e; (g) 5; (h) 7, based on their IC values. The red arrow in the
picture of cells showed the morphology of cell.

126
Z. MOKHTAR ET AL.
Figure . The molecular structure of compound 3e, drawn with % probability displacement ellipsoids
and the atom-numbering scheme. Dash lines show the C–H···O intramolecular hydrogen bonds.
Figure . (a) Crystal packing of compound 3e, hydrogen bond is shown as s dash lines. (b) Dimer in com-
pound 3e.

MOLECULAR CRYSTALS AND LIQUID CRYSTALS
127
Table . Crystal data and structure refinement of compound 3e.
CCDC
Formula

CHIO
.
Formula weight (gm·mole−
)
Temperature (K)

˚
Wavelength (A)
Crystal system
Space group
Unit cell (A, °)
Volume (A )
.
Monoclinic
P/n
˚
˚
a = .(), b = .(), c = .()β = .()

.()
Z

.
.

Calculated density (g·cm−
)
Absorption coefficient (mm−
F()
)
Crystal size (mm)
θ (min, max)
Dataset
Total/unique data/R (int)
Observed data [I > .σ(I)]
Data/parameters
R/wR/Goodness-of-fit
.×.×.
.°, .°
– ࣘ h ࣘ ; – ࣘ k ࣘ ; – ࣘ l ࣘ 
, , .

, 
., ., .
–., .
−
˚
Largest diff. peak and hole (e·A
)
These values represent the concentration of the samples that caused a 50% decrease in cell
growth [1].
The morphologic changes against MDA MB 231 and CHO cells were investigated to con-
firm the potent cytotoxicity of heterocyclic chalcones (Figures 2 and 3). It obviously showed
that the treatment of chalcones at their IC50 values caused both of the cells to be rounded in
shape, and retracted from their neighboring cells different from the non-treated cells as shown
in Figure 2(f). However, Figure 2(g) with IC50 = 23.2 µg/mL shows that the morphology of
CHO cell is the same as that of control (Figure 2a), except that the number of viable cells was
reduced (referred with red arrows).
Apart from this, it can be observed from the red arrows in Figures 2(d) and (h) of MDA
MB 231 that few of the cells still maintained their morphology when compared with their
control (Figure 3a).
Results from the data of IC50 values revealed that all chalcones possess cytotoxic activity
in a range of 9.8 to 20 µg/mL against MDA MB 231. A previous study has reported that the
compound exhibited the concentration that inhibited 50% of cell growth of CHO or MDA
231, with IC50 < 10 μg/mL considered to be as active, while IC50 > 50 μg/mL was considered
as non-cytotoxic to the cell [21]. Among all heteroarylchalcone bearing electron withdrawing
groups, Cl(3c) exhibited the highest cytotoxic activity against MDA MB 231 cell (IC50 = 9.8
μg/mL). It was supported by Solomon and Lee [4], who found that compound with 4-chloro
group attached to the phenyl ring in furan chalcone emerged as the most potent compound.
However, the introduction of halogen groups at the phenyl ring of heterocyclic chalcone,
especially compounds (3b, 3d, and 3e) caused decrease in cytotoxic activity on MDA MB 231
cancer cell than the unsubstituted chalcone (3a). Previous study showed that the introduction
of hydrophobic substituent of halogen groups (fluoro, bromo, and chloro) at the para position
of phenyl ring, especially in furan chalcone, resulted in an increase in anti-proliferative activity
than the parental compound [4].
These contradictory results have been influenced by the effect of electron affinity of the
compounds [22, 23]. According to Olive [23], a compound with higher electron affinity (abil-
ity to accept electron) required low concentration of drug. The increasing toxicity effects of
chalcone against MDA MB 231 cell by the addition of halogen groups at para position were in

128
Z. MOKHTAR ET AL.
˚
Table . Bond lengths (A), angles (◦), and torsion angles (◦) for compound 3e.
Bond lengths
I—C
. ()
. ()
. ()
. ()
. ()
. ()
.
. ()
.
. ()
. ()
.
C—C
C—C
. ()
. ()
.
. ()
.
. ()
. ()
.
. ()
.
. ()
.
.
.
O—C
O—C
O—C
C—C
C—C
C—HA
C—C
C—HA
C—C
C—C
C—HA
C—C
C—HA
C—C
C—HA
C—C
C—HA
C—C
C—C
C—HA
C—C
C—HA
C—C
C—HA
C—HB
C—HC
. ()
.
. ()
Bond angles
C—O—C
C—C—C
C—C—HA
C—C—HA
C—C—C
C—C—HA
C—C—HA
C—C—C
C—C—I
. ()
. ()
.
.
. ()
.
.
. ()
. ()
. ()
. ()
.
C—C—HA
C—C—HA
C—C—C
C—C—HA
C—C—HA
C—C—O
.
.
. ()
.
.
. ()
. ()
. ()
. ()
.
.
. ()
.
C—C—C
O—C—C
C—C—C
C—C—HA
C—C—HA
C—C—C
C—C—HA
C—C—HA
C—C—O
C—C—C
O—C—C
C—C—HA
C—C—HB
HA—C—HB
C—C—HC
HA—C—HC
HB—C—HC
C—C—I
C—C—C
C—C—HA
C—C—HA
C—C—C
C—C—HA
C—C—HA
C—C—C
C—C—C
C—C—C
O—C—C
O—C—C
C—C—C
C—C—C
.
. ()
.
.
. ()
. ()
. ()
.
.
.
.
.
.
.
. ()
. ()
. ()
. ()
. ()
. ()
. ()
Torsion angles
C—C—C—C
C—C—C—C
C—C—C—I
C—C—C—C
I—C—C—C
C—C—C—C
C—C—C—C
C—C—C—C
C—C—C—C
C—C—C—C
C—C—C—O
C—C—C—O
C—C—C—C
C—C—C—C
−.()
. ()
−.()
−.()
. ()
. ()
O—C—C—C
C—C—C—C
C—C—C—C
C—O—C—C
C—O—C—C
C—C—C—C
C—C—C—O
O—C—C—C
C—C—C—C
C—C—C—C
C—C—C—O
C—C—C—C
C—O—C—C
C—O—C—C
−.()
. ()
. ()
−. ()
−. ()
−. ()
. ()
−. ()
. ()
. ()
. ()
−. ()
−. ()
. ()
. ()
−. ()
−. ()
. ()
−. ()
. ()
. ()
−. ()
the following order: 3e < 3d < 3b < 3c. These halogen groups, which act as electron attracting
substituents, help to increase electron affinity. Conversely, literature revealed that the order of
halogen groups, as substituents were slightly different with the following order: –NO2 > –Br =
–Cl > –F [24]. This was due to several factors, such as hydrophobic parameter and molecular

MOLECULAR CRYSTALS AND LIQUID CRYSTALS
129
˚
Table . Hydrogen bond lengths (A) and angles (◦) for compound 3e.
˚
˚
˚
D–H···A
D–H (A)
H···A (A)
D···A (A)
D–H···A (◦)
C—HA···O
C—HA···Oa
C—HA···Cgb
.
.
.
.
.
.
.()
.()
.
.
.
.
Symmetry operation: (a) x, y, – + z, (b) ½ + x, ½ – y, ½ + z.
Cg: O/C–C.
hardness, that influenced the cytotoxicity of MDA MB 231 cell [22]. Meanwhile, the substitu-
tion of phenyl ring with pyrazine (5) and pyridine (7) showed weak cytotoxic effect on MDA
MB 231 cell (IC50 > 10 μg/mL). According to Ellithey et al. [25], the compound that exhibited
IC50 >10 μg/mL was considered as a weak cytotoxic to the cell.
In addition, our results clearly showed that chalcone with substituent bromine (3d) and
iodine (3e) showed greater toxicity on CHO, non-cancer cell line (13.5 and 15.0 μg/mL) com-
pared with MDA MB 231 (16.2 and 20.0 μg/mL). This SAR study implies that the toxicity of
the tested compounds on cells is possibly due to some factors such as the conjugation effect
of the presence of double bond in the structure of heterocyclic chalcone [26], the lipophilic
nature (dissolved in lipid) [4], and the presence of substituent at both aromatic and heteroaryl
rings. Overall, the synthesized compounds exert cytotoxic effect on CHO cell line, which is in
agreement with Pittella et al. [27], who reported that the chemotherapeutic agents available
in the market worked on both cancer and non-cancer cells. Apart from this, a commonly pre-
scribed anticancer agent, cisplatin, has been shown to exert cytotoxic activity on MDA MB
231 (7.10 μg/mL) and non-cancer breast cell, 184B5 (7.66 μg/mL) [4]. Survey of literature
reported that 42% of patients who were treated with high dose of cisplatin for solid tumor
suffered from nephrotoxic injury [28]. Meanwhile, tamoxifen also showed cytotoxic activity
against MDA MB 231 (12 μg/mL) and CHO cells (8 μg/mL) [5]. Based on the IC50 values
between MDA MB 231 and CHO cells, it revealed that both of these drugs are toxic toward
normal cell lines.
X-ray crystallography
All bond lengths and angles in compound 3e were within normal ranges, comparable with
the related structure [29], and listed in Table 3. The molecule exists in E configuration with
˚
respect to C8=C9 double bond (1.331(6) A). Dihedral angle between benzene and furan rings
is 12.3(3)°.
An S(5) ring motif was observed in the molecules, formed by intramolecular interaction of
C9–H9A···O2 (Table 4, and Figure 4). The crystal structure of compound 3e (Figure 5a) was
also stabilized by the intermolecular interaction of C11–H11A···O2, and formed a dimer as
shown in Figure 5b. C–H···π interaction [30] (Figure 5a) involving furan ring was also found
in the crystal structure (Table 5 ).
Conclusions
A series of heterocyclic chalcone containing halogen groups, pyrazine, and pyridine, replacing
the phenyl ring, have been successfully synthesized in high yields. The potential bioactive
compounds were evaluated for their cytotoxic activities using MTT assay. Based on the IC50
values of cytotoxic activities of heterocyclic chalcone, the overall compounds were found to
be toxic on MDA MB 231 and CHO cell as well. Since this is a preliminary study and the

130
Z. MOKHTAR ET AL.
normal cell used is from a mouse, it cannot be compared with human breast normal cell line.
In future, this series of chalcone needs to be further tested on normal breast cell lines such
as 184B5 (non-cancer breast 270 epithelial cells), as compound 3c (IC50 = 9.8 μg/mL) was
found to be an active compound against MDA MB 231.
Acknowledgments
The authors would like to thank the Tissue Engineering, Faculty of Bioscience and Medical Engineer-
ing, UTM and Biotechnology Research Laboratory 2, Faculty of Science, UTM for providing research
facilities.
Funding
The authors wish to thank the Ministry of Higher Education (MOHE) for funding this research under
the Research University Grant (RUP) with vote No. 07J48, and the Fundamental Research Grant Scheme
(FRGS) with vote No. 4F448. Hoong-Kun Fun extends his appreciation to the Deanship of Scientific
Research at King Saud University for the funding of this work through the research group project No.
RGP-VPP-207.
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