Synthesis and biological evaluation of some new triazolo[1,5-a]quinoline derivatives as anticancer and antimicrobial agents

Article abstract of DOI:10.1039/c3ra46961a

In the present study, versatile multifunctional unreported triazolo[1,5-a]quinoline derivatives were prepared. Compounds 1-19 were synthesized by adopting appropriate synthetic routes and were pharmacologically evaluated for their in vitro anticancer acti

Full text of DOI:10.1039/c3ra46961a

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Synthesis and biological evaluation of some new
triazolo[1,5-a]quinoline derivatives as anticancer
Cite this: RSC Adv., 2014, 4, 24131
and antimicrobial agents
a
Fatma A Bassyouni, Sherifa M. Abu-Baker,b Khaled Mahmoud,c Maysa Moharam,d
*
Sally S. El-Nakkadyb and Mohamed Abdel Rehime
In the present study, versatile multifunctional unreported triazolo[1,5-a]quinoline derivatives were prepared.
Compounds 1–19 were synthesized by adopting appropriate synthetic routes and were pharmacologically
evaluated for their in vitro anticancer activity against human cancer cell lines: hepatocellular liver carcinoma
(HEPG2) and Caucasian breast adenocarcinoma (MCF-7), in addition to their antibacterial and antifungal
activities. Compound 4 demonstrated strong inhibitory effects against breast cancer (MCF-7), whereas
compounds 8 and 19 exhibited moderate activity against breast carcinoma cell line MCF-7. Compounds
16 and 19 gave moderate activity against liver carcinoma cell line HEPG2. The antimicrobial activity of
the prepared compounds was tested against bacteria and fungi. Among them, the results of
antimicrobial activity indicated that compounds 4, 9, 11, 13, 15, 17, 18 and 19 were the most active
compounds. Compound 4 exhibited strong activity against Fusarium sp., whereas compounds 9, 11, 15,
17, 18 and 19 showed high activity against Escherichia coli. More specifically, compound 17 displayed a
high inhibitory effect against Bacillus cereus, Escherichia coli and Rhizoctonia sp.
Received 22nd November 2013
Accepted 7th April 2014
DOI: 10.1039/c3ra46961a
www.rsc.org/advances
pharmacological and clinical effects such as anti-viral,12 anti-
leishmanial,13 anti-tuberculosis,14 anti-inammatory,15 and
anticancer agents,16 as well as inhibitors for HIV-1 integrase.17
However, the value of synthetic quinoline-based antimalarials
has been seriously eroded in recent years, mainly as a result of
the development and spread of resistant parasites (Winstanley
and Warhurst).18,19 Chloroquine, primaquine and meoquine
have been mainstays of malaria chemotherapy, in addition to
lavendamycin, streptonigrin and streptonigrone, which have
broad antimicrobial activity.
On the other hand, coumarins, an old class of benzopyrene
compounds, are widely used in the pharmaceutical industry as
precursor molecules in the synthesis of various anti-coagulant,20
anti-HIV,21 anti-tumor22 and anti-hypertensive agents.23 These
observations aroused interest in the investigation of this
structural class, and it seemed desirable to prepare and test
some new triazolo[1,5-a]quinoline derivatives for their evalua-
tion as anticancer and antimicrobial agents because of the
aforementioned ndings and due to our interest in the
synthesis of a wide range of new heterocyclic compounds for
biological screening.24–29
1. Introduction
Cancer chemotherapy has entered a new era of molecularly
targeted therapeutics, which are highly selective and not asso-
ciated with the serious toxicities of conventional cytotoxic
drugs.1 Quinoline is a heterocyclic scaffold of paramount
importance to the human race. Several quinoline derivatives
isolated from natural resources or prepared synthetically are
signicant with respect to medicinal chemistry and biomedical
use.2,3 In addition, a series of compounds derived from quino-
line derivatives have been synthesized as potential anti-HIV
agents.4 Some quinolines showed signicant similarity to some
novel antifungal agents; therefore, they were screened for
potential antifungal activity.5,6 However, new quinoline deriva-
tives display a wide spectrum of potent herbicidal7,8 and anti-
tumor activities.9 The anti-microbial and anti-malarial activities
of quinoline derivatives have been also evaluated.10,11 Recently,
quinolines and isoquinolines have been reported to possess
aChemistry of Natural and Microbial Products Department, Pharmaceutical Research
Group, Center of Excellence for Advanced Sciences, National Research Centre,
12622, Cairo, Egypt. E-mail: fatma.nrc@hotmail.com; Fax: +20-233370931; Tel:
+20-1118596967
2. Results and discussion
bChemistry Department of Natural and Microbial Products, National Research Centre,
12622, Cairo, Egypt
The synthetic protocols of the targeted compounds are depicted
in Schemes 1–3. Coumarin derivatives are important and
fruitful starting materials used for synthesizing heterocyclic
cPharmacognosy Department, National Research Centre, 12622, Cairo, Egypt
dChemistry Department of Microbial Products, National Research Centre, 12622,
Cairo, Egypt
eDepartment of Analytical Chemistry, Stockholm University, 10691 Stockholm, Sweden compounds of pharmaceutical importance. Therefore, we are
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Scheme 1 Synthesis of 4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol derivative 3. Reagents and conditions: (i) 1 mol
semicarbazide, sodium acetate, reflux. (ii) 2 mol semicarbazide, sodium acetate, reflux. (iii) Selenium dioxide, acetic acid, reflux.
interested in synthesizing a variety of new heterocyclic triazolo corresponding to N]N. The 1H-NMR spectrum of compound 3
[1,5-a]quinoline derivatives from the coumarin moiety.
revealed the presence of multiple signals between d 6.90–7.20
Compound 1 was synthesized via the condensation reaction ppm (aromatic protons), in addition to two signals at 11.10 and
of 4-hydroxysalicylaldehyde with ethyl acetoacetate in the 11.30 ppm corresponding to two OH protons (cf. experimental
presence of 4 N HCl to afford the corresponding product iden- data).
tied as 3-acetyl-7-hydroxy-2H-chromen-2-one (1), which was
Furthermore, when compound 3 was allowed to react with
used as a starting material. The synthesis of this product was formaldehyde and amino acids, such as D-glycine, D-alanine, D-
accomplished through a straightforward adaptation of the phenylalanine, D-histidine, and 2-aminobutyric acid in the
previously reported synthesis by Furniss et al. and Singh presence of absolute ethanol and TEA, it led to the formation of
et al.30,31 The reaction of compound 1 with one mole of semi- the corresponding 2-((2,8-dihydroxy-4-(6H-1,2,3-selenadiazol-4-
carbazide in the presence of sodium acetate32 led to the yl)-[1,2,4]triazolo[1,5-a]quinolin-9-yl)methylamino) carboxylic
formation of 1-(2,8-dihydroxy-[1,2,4]triazolo[1,5-a]quinolin-4-yl)- acid derivatives (4–8) (Scheme 2). The structures of compounds
ethanone (2a). The structure of compound 2a was conrmed by 4–8 were established on the basis of elemental analysis and
spectroscopic data and elemental analysis. The IR spectrum of spectral data (cf. experimental data). As an example, the IR
product 2a indicated the presence of two bands (OH groups) at spectrum of compound 4 showed absorption bands at 3335–
3400 and 3430 cmꢀ1, one band (C]O group) at 1690 cmꢀ1, in 3450 cmꢀ1 (two OH groups), another absorption band at 1715
addition to two bands at 1640 and 1645 cmꢀ1 due to the pres- cmꢀ1 (C]O acid), in addition to a band at 1640 cmꢀ1 (C]N).
1
ence of two C]N groups. The 1H-NMR spectrum of compound The H-NMR spectrum of compound 4 indicated the presence
2a showed two OH protons at d 11.15 and 11.40 ppm, in addi- of signals at d 11.00–11.40 ppm for OH protons, an NH proton at
tion to aromatic protons at d 6.90–7.25 ppm and disappearance d 9.80 ppm and aromatic protons between d 6.80–7.00 ppm. The
of the lactone band. The mass spectrum of 2a showed a IR spectrum of compound 5 exhibited absorption bands at
compound with m/z at 243 [M+].
3448–3458 cmꢀ1 (attributed to the OH groups), 1700 cmꢀ1
The target product 2a reacted with another one mole of (corresponding to C]O acid) and 1570 cmꢀ1 (for N]N group).
semicarbazide in sodium acetate to afford the corresponding
The 1H-NMR spectrum of compound 6 showed the presence
product 2-(1-(2,8-dihydroxy-[1,2,4]triazolo[1,5-a]quinolin-4-yl)- of signals between d 6.86–7.45 ppm for aromatic protons, at d
ethylidene)hydrazinecarboxamide (2b). The IR spectrum of 9.40 ppm for NH proton, and at d 11.10, 11.28 and 11.50 ppm for
compound 2b indicated the presence of two OH groups at 3445 OH protons.
and 3455 cmꢀ1, an NH absorption band at 3340 cmꢀ1 and an
The formation of 9-(1-(3H-imidazo[4,5-b]pyridin-2-yl)-4-
absorption band at 3286 cmꢀ1 (NH2). In addition, the IR spec- (1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol)
trum showed a C]O amide band at 1688 cmꢀ1 and two derivatives 9–13 was achieved via condensation and cyclization
absorption bands at 1635 and 1645 cmꢀ1 (C]N groups), in reactions of the open compounds 4–8 with 2,3-diaminopyridine
addition to a peak in the mass spectrum with m/z at 300 [M+].
in the presence of anhydrous pyridine (cf. Scheme 2). The
The use of selenium dioxide as an oxidative agent is very well structures of the synthesized compounds 9–13 were conrmed
documented.33,34 The oxidative cyclization of compound 2b with by their corrected elemental analyses as well as spectral data. As
selenium dioxide using acetic acid gave the corresponding 4-(6H- an example, compound 9 can be structurally elucidated from
1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol analytical analysis and spectral data as follows: the 1H NMR
(3). The IR spectrum of compound 3 revealed absorption bands spectrum showed signals at d 3.95 ppm, 4.20 ppm for two CH2
at 3435 and 3445 cmꢀ1 (two OH groups), 1635 and 1644 cmꢀ1 protons, d 6.80–7.10 ppm for aromatic protons, at d 9.70 and
(C]N groups), in addition to an absorption band at 1575 cmꢀ1 10.80 ppm for NH protons, and d 11.40 and 11.58 ppm for OH
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Scheme
2
Synthesis of 2-((2,8-dihydroxy-4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinolin-9-yl)methylamino) carboxylic acid
derivatives (4–8) and 9-((1-(3H-imidazo[4,5-b]pyridin-2-yl)-4-(1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol derivatives (9–
13). Reagents and conditions: amino acids, i.e., D-glycine, D-alanine, D-phenylalanine, D-histidine, or 2-aminobutyric acid; formaldehyde;
absolute ethanol; TEA and reflux at 60 ꢁC. (i–v) 2,3-Diaminopyridine, dry pyridine, stirring at room temperature for 1 h, then reflux.
protons. The IR spectrum displayed an absorption band at 3420–3450 cmꢀ1 (OH groups), 3380–3395 cmꢀ1 (NH groups), in
3335–3359 cmꢀ1 for NH groups and at 3445–3460 cmꢀ1 for OH addition to absorption bands at 1710–1725 cmꢀ1 (C]O acid)
groups (cf. experimental data).
and at 1640 and 1645 cmꢀ1 (C]N group).
The IR spectrum of compound 10 showed absorption bands
Finally, compounds 14–16 were readily condensed and
at 3438 and 3447 cmꢀ1 (2 OH), 3332 and 3348 cmꢀ1 (2 NH), 1640 cyclized to the corresponding 2-(2,8-dihydroxy-4-acetyl-[1,2,4]
cmꢀ1 (C]N) and 1570 cmꢀ1 (N]N).
triazolo[1,5-a]quinolin-9-yl)methylamino-9-(benzimidazole-
1
The H NMR spectrum of compound 13 revealed the pres- carboxylic acid) derivatives (17–19) upon treatment with 3,4-
ence of multiple signals between d 6.86–7.45 ppm for aromatic diaminobenzoic acid in the presence of anhydrous pyridine (cf.
protons and two signals at 9.50 and 10.78 ppm for two NH Scheme 3). The structures of compounds 17–19 were deduced
protons, in addition to two bands at 11.45 and 11.56 ppm for from their elemental analyses and spectral data.
two OH protons (cf. experimental data).
The IR spectra of compounds 17–19 indicated the presence of
When compound 2a reacted with amino acids such as D- OH absorption bands at 3415–3465 cmꢀ1, NH absorption bands
tyrosine, D-phenylalanine or D-histidine and formaldehyde in at 3370–3395 cmꢀ1, and CO carboxylic acid at 1710–1728 cmꢀ1
the presence of absolute ethanol and TEA under reux condi-
.
tions, it gave the corresponding 2-((2,8-dihydroxy-4-acetyl-[1,2,4]-
triazolo[1,5-a]quinolin-9-yl)methylamino) carboxylic acid deriv-
atives (14–16) (Scheme 3). The structures of compounds 14–16
3. Experimental
3.1 General
were conrmed by their spectral data and elemental analyses. Reagents used for synthesis were purchased from Sigma-
The IR spectra displayed characteristic absorption bands at Aldrich and Merck. All melting points were determined using
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Scheme 3 Synthesis of 2-((2,8-dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]quinolin-9-yl)methylamino) carboxylic acid derivatives (14–16) and 2-
((2,8-dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]quinolin-9-yl)methylamino-9-(benzimidazole-carboxylic acid) derivatives (17–19). Reagents and
conditions: (i) semicarbazide, absolute ethanol, TEA, reflux; (ii) D-tyrosine, HCHO, absolute ethanol, reflux; (iii) D-phenylalanine, HCHO, absolute
ethanol, TEA, reflux; (iv) D-histidine, HCHO, absolute ethanol, TEA, reflux at 60 ꢁC; (v–vii) 3,4-diaminobenzoic acid, anhydrous pyridine, reflux.
an electrothermal capillary melting point apparatus (Stuart,
SMP30, UK) and are uncorrected. 1H NMR and 13C NMR spectra
were measured in DMSO-d6 using a JEOL-300 spectrometer
(USA), and chemical shis were expressed as ppm against TMS
as an internal reference. Mass spectra were obtained using a
GC-MS Finnigan spectrometer (USA) at 70 eV. The IR spectra
(4000–400 cmꢀ1) were recorded from thin lms using KBr
pellets in a Jasco FT/IR 300 E Fourier transform infrared spec-
trophotometer (JASCO Corporation, Tokyo, Japan). Elemental
analyses were performed at the micro analytical laboratory of
Cairo University, Cairo, Egypt. The monitoring of the progress
of all reactions and homogeneity of the synthesized compounds
was carried out by TLC (silica gel pre-coated aluminum cards
with uorescent indicator at 254 nm, Merck, Germany). Visu-
alization was performed by illumination with a UV light source.
3.2 Synthesis
Synthesis of 3-acetyl-7-hydroxy-2H-chromen-2-one (1) was per-
formed by the modied procedure of Furniss et al. and Singh
et al.30,31
Synthesis of 1-(2,8-dihydroxy-[1,2,4]triazolo[1,5-a]quinolin-4-
yl)ethanone (2a) and 2-(1-(2,8-dihydroxy-[1,2,4]triazolo[1,5-a]-
quinolin-4-yl)ethylidene)hydrazinecarboxamide (2b)
General procedure. A solution of semicarbazide (0.01 mol) and
crystalline sodium acetate (0.02 mol) in water (10 ml) was added
with stirring to a solution of compound 1 (0.01 mol) in absolute
ethanol (50 ml) containing a catalytic amount of glacial acetic
acid (1 ml). Stirring was continued for 4–5 h. The reaction
mixture was poured onto crushed ice and then was cooled in a
refrigerator overnight. During the procedure the reaction
progress was monitored by TLC (eluent: n-hexane–ethyl acetate:
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2 : 1). The precipitated solid was ltered, washed with cold with stirring for 5–6 h. The completion of the reaction was
water, dried under vacuum and then crystallized from absolute conrmed by TLC (eluent: n-hexane–ethyl acetate: 2 : 1). The
ethanol to afford compound 2a.
product was then cooled to room temperature. The solvent was
1-(2,8-Dihydroxy-[1,2,4] triazolo[1,5-a]quinolin-4-yl)ethanone evaporated under vacuum and the precipitated solid obtained
(2a). Yield: 75%. m.p. 198–200 ꢁC. 1H-NMR: (300 MHz, DMSO): d was dried to afford compounds 4–8.
2.26 (3H, s, CH3), 6.90–7.25 (4H, m, Ar-H aromatic proton),
2-((2,8-Dihydroxy-4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo-
11.15 (1H, br s, OH-triazole), 11.40 (1H, br s, Ar-OH). 13C NMR [1,5-a]quinolin-9-yl)methylamino)acetic acid (4). Yield 72%; m.p.
(DMSO-d6): d 20.90 (CH3), 110.90, 115.80, 117.90, 119.80, 180–182 ꢁC; 1H-NMR (300 MHz, DMSO-d6): d 4.30 (2H, s, CH2–),
122.90, 127.00, 130.00, 135.80, 165.90, 168.00 (C]N), 171.60 4.50 (2H, d, CH2–), 5.80 (1H, s, CH selenadiazol proton), 6.80–
(C]O). IR (KBr): n ¼ 3400, 3430 (2OH), 1690 (CO), 1640, 1645 7.00 (3H, m, Ar-H), 9.80 (1H, br s, NH–, D2O-exchangeable), 11.00
(2C]N) cmꢀ1. MS: m/z ¼ 243 [M+]; anal. calcd for C12H9N3O3: C, (1H, br s, COOH), 11.20 (1H, br s, OH-triazole proton), 11.40 (1H,
59.25; H, 3.70; N, 17.28; found: C, 59.29; H, 3.73; N, 17.24.
br s, OH-aromatic proton); 13C NMR (DMSO-d6): d 39.90 (CH2),
2-(1-(2,8-Dihydroxy-[1,2,4]triazolo[1,5-a]quinolin-4-yl)ethyl- 41.90 (CH2), 110.60, 113.90, 118.50, 119.60, 121.80, 125.20,
idene)hydrazinecarboxamide (2b) was prepared according to 127.90, 131.50, 134.80, 136.90, 156.60, 159.80 (C]N), 178.80
the same procedure as 2a.
(CO acid); IR (KBr) n ¼ 3420, 3435, 3450 (3 OH), 3168 (CH
2-(1-(2,8-dihydroxy-[1,2,4]triazolo[1,5-a]quinolin-4-yl)ethylidene)- aromatic), 1715 (CO carboxylic acid), 1640 (C]N), 1630 (C]C),
hydrazinecarboxamide (2b). Yield 78%; m.p. 170–172 ꢁC; 1H- 1570 (N]N) cmꢀ1; anal. calcd for C15H12N6O4Se: C, 42.97; H,
NMR (300 MHz, DMSO-d6): d 2.18 (3H, s, CH3), 6.40 (2H, s, NH2), 2.88; N, 20.05; Se, 18.83; found: C, 43.07; H, 2.94; N, 20.10; Se,
6.80–7.00 (4H, m, Ar-H aromatic proton), 10.30 (1H, br s, NH, 18.87.
D2O-exchangeable), 11.25 (1H, br s, OH-triazole proton), 11.45
2-((2,8-Dihydroxy-4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo-
(1H, br s, OH-aromatic proton), 13C NMR (DMSO-d6): d 22.10 [1,5-a]quinolin-9-yl)methylamino)propanoic acid (5). Yield 70%;
1
ꢁ
(CH3), 110.80, 111.90, 115.80, 117.90, 118.80, 122.59, 130.20, m.p. 190–192 C; H-NMR (300 MHz, DMSO-d6): d 2.20 (3H, d,
134.60, 161.90, 166.20 (C]N), 175.60 (C]O); IR (KBr) n ¼ 3440, CH3–), 4.10 (2H, s, CH2–), 5.65 (1H, s, CH selenadiazol proton),
3445 (2 OH), 3340 (NH), 3286 (NH2), 3165 (CH aromatic), 1688 5.99 (1H, q, CH–), 6.90–7.10 (3H, m, Ar-H), 9.50 (1H, br s, NH–
(C]O amide), 1635, 1645 (2C]N), 1630 (C]C) cmꢀ1; MS: m/z CH, D2O-exchangeable), 11.20 (1H, br s, COOH), 11.40 (1H, br s,
¼ 300 [M+]; anal. calcd for C13H12N6O3. C, 52.00; H, 4.03; N, OH-triazole), 11.60 (1H, br s, OH-aromatic); 13C NMR (DMSO-
27.99; found C, 52.08; H, 4.10; N, 28.06.
d6): d 24.90 (CH3), 40.90 (CH2), 50.60 (CH), 111.60, 114.90,
Synthesis of 4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]- 116.50, 118.60, 121.80, 124.50, 126.80, 131.50, 134.80, 136.90,
quinoline-2,8-diol (3). A solution of compound 2b (5 mmol) in 158.60, 161.80 (C]N), 168.80 (CO acid); IR (KBr) n ¼ 3430, 3440,
glacial acetic acid (15 ml) was warmed to 80 ꢁC with stirring, 3455 (3 OH), 3165 (CH aromatic), 1700 (CO carboxylic acid),
then selenium dioxide (0.55 g, 5 mmol) was added dropwise 1658 (C]N), 1635 (C]C), 1570 (N]N) cmꢀ1; anal. calcd for
during a period of 30 min and stirring was continued for a
C16H14N6O4Se: C, 44.35; H, 3.26; N, 19.40; O, 14.77; Se, 18.22;
further 4 h (TLC control). Aer completion of the reaction, the found: C, 44.39; H, 3.32; N, 19.48; Se, 18.27.
reaction mixture was ltered to remove the deposited selenium.
2-((2,8-Dihydroxy-4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo-
The ltrate was poured onto crushed ice and the obtained solid [1,5-a]quinolin-9-yl)methylamino)-3-phenylpropanoic acid (6).
1
ꢁ
was ltered off and washed thoroughly with cold water, sodium Yield 71%; m.p. 150–152 C; H-NMR (300 MHz, DMSO-d6): d
carbonate solution and water (10%). The obtained product aer 3.82 (2H, d, CH2), 4.30 (2H, d, CH2–), 5.70 (1H, s, CH selena-
drying was recrystallized from absolute ethanol to give diazol proton), 5.95 (2H, t, CH2–), 6.86–7.12 (3H, m, Ar-H
compound 3.
aromatic proton), 7.29–7.45 (5H, m, Ar-H), 9.40 (1H, br s, NH,
4-(6H-1,2,3-Selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline- D2O-exchangeable), 11.10 (1H, br s, COOH), 11.28 (1H, br s, OH-
2,8-diol (3). Yield 74%; m.p. 200–202 ꢁC; 1H-NMR (300 MHz, triazole), 11.50 (1H, br s, OH aromatic); IR (KBr) n ¼ 3415, 3430,
DMSO-d6): d 5.90 (1H, s, CH-selenadiazol), 6.90–7.20 (4H, m, Ar- 3440 (3 OH), 3169 (CH aromatic), 1715 (CO carboxylic acid),
H aromatic proton), 11.10 (1H, br s, OH-triazole proton), 11.30 1650 (C]N), 1636 (C]C), 1575 (N]N) cmꢀ1; anal. calcd for
(1H, br s, OH-aromatic proton); 13C NMR (DMSO-d6): d 114.60,
116.90, 117.90, 118.90, 122.60, 124.50, 126.90, 131.50, 135.80, 51.80; H, 3.6; N, 16.55; O; Se, 15.56.
C22H18N6O4Se: C, 51.87; H, 3.56; N, 16.50; Se, 15.50; found: C,
136.90, 156.30, 158.90 (C]N); IR (KBr) n ¼ 3435, 3445 (2 OH),
2-((2,8-Dihydroxy-4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo-
3170 (CH aromatic), 1635, 1644 (C]N), 1633 (C]C), 1575 (N] [1,5-a]quinolin-9-yl)methylamino)-3-(1H-imidazol-4-yl)propanoic
N) cmꢀ1; MS: m/z ¼ 332 [M+ ꢀ 1]; anal. calcd for C12H7N5O2Se: acid (7). Yield 71%; m.p. 140–142 ꢁC; 1H-NMR (300 MHz, DMSO-
C, 43.38; H, 2.10; N, 21.09; Se, 23.77; found: C, 34.34; H, 2.14; N, d6): d 3.85 (2H, s, CH2–), 4.15 (2H, d, CH2–), 5.60 (1H, s, CH
21.02; Se, 23.72.
selenadiazol proton), 5.90 (1H, t, CH–), 6.10 (1H, s, –CH–NH
Synthesis of 2-((2,8-dihydroxy-4-(6H-1,2,3-selenadiazol-4-yl)- imidazole proton), 6.60 (1H, s, CH-imidazole proton), 6.90–7.18
[1,2,4]triazolo[1,5-a]quinolin-9-yl)methylamino) carboxylic acid (3H, m, Ar-H), 9.60 (1H, br s, CH2–NH, D2O-exchangeable),
derivatives (4–8)
10.40 (1H, br s, NH, imidazole D2O-exchangeable), 11.20 (1H, br
General procedure. A mixture of compound 3, amino acids s, COOH), 11.50 (1H, br s, OH-triazole), 11.70 (1H, s, OH-
(0.05 mmol), i.e., D-glycine, D-alanine, D-phenylalanine, D-histi- aromatic); IR (KBr) n ¼ 3420, 3437, 3446 (3 OH), 3164 (CH
dine, or 2-aminobutyric acid, and formaldehyde (1.25 mmol)ꢁin aromatic), 1710 (CO carboxylic acid), 1658 (C]N), 1633 (C]C),
absolute ethanol (30 ml) containing TEA was reuxed at 60 C 1573 (N]N) cmꢀ1; anal. calcd for C19H16N8O4Se: C, 45.70; H,
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(11). Yield 72%; m.p. 124–126 ꢁC. 1H-NMR (300 MHz, DMSO-d6):
d 3.90 (2H, s, CH2–NH), 4.15 (2H, d, –CH–CH2), 5.58 (1H, t, –CH–),
5.70 (1H, s, CH-selenadiazol proton), 6.75–7.00 (3H, m, Ar-H
aromatic proton), 7.10–7.25 (5H, m, Ar-H aromatic proton), 7.35–
7.55 (3H, m, Ar-H-pyridine), 9.68 (1H, br s, NH, D2O-exchange-
able), 10.50 (1H, br s, NH, D2O-exchangeable), 11.25 (1H, s, OH-
triazole), 11.45 (1H, s, OH-aromatic); IR (KBr) n ¼ 3445, 3455 (2
OH), 3356, 3368 (2 NH), 1649 (C]N), 1635 (C]C), 1572 (N]N)
cmꢀ1; MS: m/z ¼ 581 [M+ ꢀ 1]; anal. calcd for C27H21N9O2Se: C,
55.67; H, 3.63; N, 21.64; O, 5.49; Se, 13.56; found: C, 55.72; H,
3.68; N, 21.64; Se, 13.62.
9-((1-(3H-Imidazo[4,5-b]pyridin-2-yl)-2-(1H-imidazol-4-yl)ethyl-
amino)methyl)-4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]-
quinoline-2,8-diol (12). Yield 73%; m.p. 184–186 ꢁC. 1H-NMR
(300 MHz, DMSO-d6): d 3.90 (2H, s, CH2–), 4.15 (2H, d, CH2–),
5.40 (1H, t, –CH–), 5.60 (1H, s, CH-selenadiazol), 5.90 (1H, s,
–CH-imidazole proton), 6.10 (1H, s, –CH-imidazole proton),
6.80–7.15 (3H, m, Ar-H aromatic proton), 7.29–7.58 (3H, m, Ar-
H), 9.50 (1H, br s, NH-imidazole, D2O-exchangeable), 9.69 (1H,
br s, NH, D2O-exchangeable), 10.58 (1H, br s, NH, D2O-
exchangeable), 11.27 (1H, br s, OH-triazole), 11.48 (1H, br s, OH-
aromatic proton); IR (KBr) n ¼ 3433, 3445 (2 OH), 3348, 3360 (2
NH), 1650 (C]N), 1635 (C]C), 1574 (N]N) cmꢀ1; MS: m/z ¼
570 [M+ ꢀ 2]; anal. calcd for C24H19N11O2Se: C, 50.36; H, 3.35; N,
26.92; O, 5.59; Se, 13.79; found: C, 50.31; H, 3.39; N, 26.99; Se,
13.85.
9-((1-(3H-Imidazo[4,5-b]pyridin-2-yl)propylamino)methyl)-4-
(1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol
(13). Yield 73%; m.p. 160–160 ꢁC. 1H-NMR (300 MHz, DMSO-d6):
d 1.38 (3H, t, CH3), 3.85 (2H, s, CH2–), 4.28 (2H, m, CH2–), 5.55
(1H, t, CH–), 5.77 (1H, s, CH-selenadiazol), 6.86–7.20 (3H, m, Ar-
H aromatic proton), 7.30–7.45 (3H, m, Ar-H-pyridine proton),
9.50 (1H, br s, NH, D2O-exchangeable), 10.78 (1H, br s, NH, D2O-
exchangeable), 11.45 (1H, br s, OH-triazole), 11.56 (1H, br s, OH
aromatic); IR (KBr) n ¼ 3446, 3458 (2 OH), 3352, 3368 (2 NH),
1659 (C]N), 1638 (C]C), 1578 (N]N) cmꢀ1; MS: m/z ¼ 518 [M+
ꢀ 2]; anal. calcd for C22H19N9O2Se: C, 50.77; H, 3.68; N, 24.22; O,
6.15; Se, 15.17; found: C, 50.72; H, 3.74; N, 24.25; Se, 15.23.
Synthesis of 2-(2,8-dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]-
quinolin-9-yl) carboxylic acid derivatives (14–16)
3.23; N, 22.44; O, 12.82; Se, 15.81; found: C, 45.78; H, 3.29; N,
22.50; Se, 15.88.
2-((2,8-Dihydroxy-4-(6H-1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo-
[1,5-a]quinolin-9-yl)methylamino)butanoic acid (8). Yield 68%;
ꢁ
m.p. 130–132 C; 1H-NMR (300 MHz, DMSO-d6): d 2.18 (3H, t,
CH3), 3.85 (2H, s, CH2–), 4.15 (2H, m, CH2–), 5.55 (1H, t, CH–),
5.70 (1H, s, CH selenadiazol proton), 6.80–7.10 (3H, m, Ar-H),
9.20 (1H, br s, NH–, D2O-exchangeable), 11.15 (1H, br s, OH),
11.35 (1H, br s, OH-triazole), 11.58 (1H, br s, OH aromatic); 13
C
NMR (DMSO-d6): d 20.75 (CH3), 39.10 (CH2), 42.19 (CH2), 51.80
(CH), 112.60, 113.80, 117.50, 118.90, 119.60, 124.80, 125.20,
126.90, 131.50, 134.80, 136.90, 152.60, 159.30 (C]N), 170.80
(CO acid); IR (KBr) n ¼ 3415, 3430, 3440 (3 OH), 3168 (CH
aromatic), 1720 (CO carboxylic acid), 1650 (C]N), 1638 (C]C),
1578 (N]N) cmꢀ1; anal. calcd for C17H16N6O4Se: C, 45.65; H,
3.61; N, 18.79; Se, 17.60; found: C, 45.72; H, 3.68; N, 18.84; Se,
17.69.
Synthesis of 9-(1-(3H-imidazo[4,5-b]pyridin-2-yl)-4-(1,2,3-
selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol) deriv-
atives (9–13)
General procedure. A mixture of compounds 4–8 (0.01 mmol)
and 2,3-diaminopyridine (0.01 mmol) was dissolved in dry
pyridine (10 ml) with stirring at room temperature for 1 h, and
then heated under reux for 5–6 h. The completion of the
reaction was conrmed by TLC (eluent: petroleum ether (60–
80)–ethyl acetate: 9 : 1). The reaction was cooled to room
temperature, and then poured into crushed ice containing HCl
(2 ml of 10% HCl). The formed precipitate was allowed to settle
down overnight at room temperature, and then ltered, dried
under vacuum and recrystallized from absolute ethanol to
produce compounds 9–13.
9-(((3H-Imidazo[4,5-b]pyridin-2-yl)methylamino)methyl)-4-
(1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol
(9). Yield 74%; m.p. 166–186 ꢁC. 1H-NMR (300 MHz, DMSO-d6):
d 3.95 (2H, s, CH2–), 4.20 (2H, s, CH2–), 5.86 (1H, s, CH-selena-
diazol proton), 6.80–7.12 (3H, m, Ar-H aromatic), 7.20–7.48 (3H,
m, Ar-H-pyridine), 9.70 (1H, br s, NH, D2O-exchangeable), 10.80
(1H, br s, NH, D2O-exchangeable), 11.42 (1H, br s, OH-triazole),
11.58 (1H, br s, OH-aromatic); IR (KBr) n ¼ 3445, 3460 (2 OH),
3335, 3359 (2 NH), 1654 (C]N), 1630 (C]C), 1576 (N]N) cmꢀ1
;
MS: m/z ¼ 491[M+ ꢀ 1]; anal. calcd for C20H15N9O2Se: C, 48.79;
H, 3.07; N, 25.60; Se, 16.04; found: C, 48.70; H, 3.15; N, 25.69; O;
Se, 16.10.
General procedure. A mixture of 1-(2,8-dihydroxy-[1,2,4]tri-
azolo[1,5-a]quinolin-4-yl)ethanone 2a (0.01 mol), an amino
acid, i.e., D-tyrosine, D-phenylalanine or D-histidine (0.01 mol),
and formaldehyde (0.01 mol) in absolute ethanol (30 ml) con-
taining triethylamine (TEA, 1 ml) was reuxed with stirring for
5–6 h at 60 ꢁC. The reaction mixture was monitored by TLC
(eluent: n-hexane–ethyl acetate: 2 : 1). The solvent was then
evaporated under vacuum, and the precipitate obtained was
ltered and dried under vacuum to afford compounds 14–16.
2-((2,8-Dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]quinolin-9-yl)-
methylamino)-3-(4-hydroxyphenyl)propanoic acid (14). Yield:
9-((1-(3H-Imidazo[4,5-b]pyridin-2-yl)ethylamino)methyl)-4-
(1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol
(10). Yield 75%; m.p. 177–179 ꢁC. 1H-NMR (500 MHz, DMSO-d6):
d 2.18 (3H, s, CH3), 4.25 (2H, s, CH2–), 5.20 (1H, q, –CH–), 5.65
(1H, s, CH-selenadiazol), 6.90–7.18 (3H, m, Ar-H aromatic
proton), 7.15–7.32 (3H, m, Ar-H-pyridine), 9.60 (1H, br s, NH,
D2O-exchangeable), 10.55 (1H, br s, NH, D2O-exchangeable),
11.20 (1H, br s, OH-triazole), 11.40 (1H, br s, OH-aromatic); IR
(KBr) n ¼ 3438, 3447 (2 OH), 3332, 3348 (2 NH), 1640 (C]N),
1633 (C]C), 1570 (N]N) cmꢀ1; MS: m/z ¼ 505 [M+ ꢀ 1]; anal.
calcd for C21H17N9O2Se: C, 49.81; H, 3.38; N, 24.89; Se, 15.59;
found: C, 49.86; H, 3.32; N, 24.84; Se, 15.53.
1
ꢁ
65%; m.p. 86–90 C. H-NMR (300 MHz, DMSO): d 2.18 (3H, s,
CH3), 4.10 (2H, d, CH2–), 4.28 (2H, s, CH2–), 5.60 (1H, t, CH–),
6.70–6.95 (3H, m, Ar-H), 7.30–7.50 (4H, m, Ar-H), 9.30 (1H, br s,
NH–), 11.25 (1H, br s, C6H4–OH), 11.46 (1H, br s, COOH), 11.55
(1H, br s, OH-triazole), 11.65 (1H, br s, Ar-OH); IR (KBr) n ¼
9-((1-(3H-Imidazo[4,5-b]pyridin-2-yl)-2-phenylethylamino)methyl)-
4-(1,2,3-selenadiazol-4-yl)-[1,2,4]triazolo[1,5-a]quinoline-2,8-diol
24136 | RSC Adv., 2014, 4, 24131–24141
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3420–3450 (4 OH), 3390 (NH), 1710 (CO carboxylic acid), 1640, triazole), 11.60 (1H, br s, Ar-OH); IR (KBr) n ¼ 3430, 3440, 3455 (3
1645 (C]N) cmꢀ1; MS: m/z ¼ 236 [M+, 36%]; anal. calcd for OH), 3380, 3390 (2 NH), 1720 (CO carboxylic acid), 1649, 1645,
C
22H20N4O6: C, 60.55; H, 4.58; N, 12.84; found: C, 60.50; H, 4.52; 1640 (3C]N) cmꢀ1. MS: m/z ¼ 534 [M+ ꢀ 2, 21%]; anal. calcd for
N, 12.76.
C29H24N6O5: C, 64.92; H, 4.47; N, 15.67; found: C, 64.85; H, 4.42;
2((2,8-Dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]quinolin-9-yl)- N, 15.61.
methylamino)-3-phenylpropanoic acid (15). Yield: 67%; m.p. 160–
2-((2,8-Dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]quinolin-9-yl)-
162 ꢁC. 1H-NMR (300 MHz, DMSO): d 2.18 (3H, s, CH3), 4.35 (2H, methylamino)-3-(1H-imidazol-4-yl)-9-(1H-benzoimidazole-carbox-
d, CH2–), 4.48 (2H, s, CH2–), 5.58 (1H, t, CH–), 6.75–6.97 (3H, m, ylic acid) (19). Yield 63%; m.p. 173–175 ꢁC. 1H-NMR (300 MHz,
Ar-H), 7.40–7.65 (5H, m, Ar-H), 9.45 (1H, br s, NH–), 10.99 (1H, DMSO): d 2.19 (3H, s, CH3), 4.10 (2H, d, CH2–), 4.45 (2H, s, CH2–
br s, COOH), 11.30 (1H, br s, OH-triazole), 11.60 (1H, s, Ar-OH); ), 5.83 (1H, t, CH–), 6.45 (1H, s, CH-imidazole), 6.85–7.20 (3H, m,
IR (KBr) n ¼ 3395 (NH), 3410, 3430, 3450 (3 OH), 1715 (CO Ar-H), 7.30–7.50 (3H, m, Ar-H), 8.80 (1H, s, CH]N-imidazole),
carboxylic acid), 1630, 1640 (2C]N) cmꢀ1; anal. calcd for C22 9.57 (1H, br s, NH–), 10.90 (1H, br s, COOH), 11.40 (1H, br s, OH-
H20N4O5: C, 62.85; H, 4.76; N, 13.33; found: C, 62.78; H, 4.71; N, triazole), 11.65 (1H, br s, NH-benzimidazole), 11.80 (1H, br s, Ar-
13.36.
OH), 11.90 (s, 1H, NH-imidazole); IR (KBr) n ¼ 3415, 3430, 3445,
2-((2,8-Dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a] quinolin-9-yl)- (3 OH), 3370, 3380, 3395 (3 NH), 1710 (CO carboxylic acid), 1635,
methylamino)-3-(1H-imidazol-4-yl)propanoic acid (16). Yield: 1640, 1644, 1648 (C]N), 1240 (C]C) cmꢀ1. MS: m/z ¼ 524 [M+
64%; m.p. 233–235 ꢁC 1H-NMR (300 MHz, DMSO): d 2.35 (3H, s, ꢀ 2]; anal. calcd for C26H22N8O5: C, 59.31; H, 4.18; N, 21.29; O,
CH3), 4.20 (2H, d, CH2–), 4.35 (2H, d, CH2–), 4.55 (1H, m, CH–), 15.21; found: C, 59.28; H, 4.15; N, 21.30; O, 15.22.
5.80 (1H, s, CH–NH-imidazole), 7.15–7.38 (3H, m, Ar-H), 8.78
(1H, s, CH]N-imidazole), 9.50 (1H, br s, NH–), 10.90 (1H, br s,
COOH), 11.30 (1H, br s, OH-triazole), 11.50 (1H, br s, Ar-OH),
4. Anticancer activity
4.1 Cytotoxic effect on human cell lines HEPG2 and MCF7
11.80 (1H, br s, NH-imidazole); IR (KBr) n ¼ 3415, 3430, 3445, (3
OH), 3385, 3395 (2 NH), 1718 (CO carboxylic acid), 1645, 1640,
The antitumor activities of the synthesized compounds were
1630 (3C]N), 1250 (C]C) cmꢀ1; anal. calcd for C19H18N6O5: C,
tested for their cytotoxic activity against two different cell lines:
55.60; H, 4.39; N, 20.48; found: C, 55.66; H, 4.32; N, 20.41.
a liver carcinoma cell line (HEPG2) and a human breast carci-
Synthesis of 2-(2,8-dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]-
noma cell line (MCF-7). The antitumor activities were evaluated
using doxorubicin as a reference cytotoxic drug for both cell
quinolin-9-yl)methylamino-9-(1H-benzimidazole-carboxylic
acid) derivatives (17–19)
General procedure. A mixture of compounds 14, 15 or 16 (0.01
mol) and 1,2-diamino-4-benzoic acid (0.01 mol) in dry pyridine
lines. The most promising compounds that gave 60% cytotox-
icity or more at a concentration of 100 mg mlꢀ1 were subjected
to comprehensive evaluation to calculate their IC50 values
(10 ml) was stirred at room temperature for about 30 minutes.
(Tables 1 and 2). Cell culture cytotoxicity assays were carried out
ꢁ
The solution was then heated under reux for 5–6 h at 80 C.
as described previously.35
The completion of the reaction was conrmed by TLC (eluent:
Studies concerning the cytotoxicity effect towards the HEPG2
petroleum ether (60–80)–ethyl acetate: 2 : 1). The reaction
cell line compared to the reference drug doxorubicin showed
that compounds 16 and 19 exhibited moderate activity. Results
for the cytotoxicity effect towards the human breast carcinoma
cell line MCF-7 compared to the reference drug doxorubicin
indicated that compound 4 was the most potent compound
tested, while compounds 19 and 8 exhibited moderate activity,
and compounds 17, 12 and 7 exhibited a weak cytotoxic effect in
both cell lines.
mixture was then cooled to room temperature and poured into
crushed ice, neutralized by hydrochloric acid (10%). The
product was collected by ltration, dried and recrystallized from
ethanol to give the desired products 17–19.
2-((2,8-Dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]quinolin-9-yl)-
methylamino)-3-(1H-4-methyl-benzo-4-yl)-9-(1H-benzoimidazole-
carboxylic acid) (17). Yield: 68%; m.p. 138–141 ꢁC. 1H-NMR (300
MHz, DMSO): d 2.25 (3H, s, CH3), 4.20 (2H, d, CH2–), 4.45 (2H, s,
CH2–), 5.58 (1H, t, CH–), 6.79–6.95 (3H, m, Ar-H), 7.10–7.30 (3H,
m, Ar-H), 7.36–7.55 (4H, m, Ar-H), 9.38 (1H, br s, NH–), 9.80 (1H,
4.2 Biological screening: in vitro assay for anticancer activity
br s, NH-benzimidazole), 10.90 (1H, br s, Ar–OH), 11.10 (1H, br The synthesized compounds were supplied to the Bioassay-Cell
s, COOH), 11.30 (1H, br s, OH-triazole), 11.70 (1H, s, Ar-OH); IR Culture Laboratory, National Research Center, Cairo, Egypt, for
(KBr) n ¼ 3425, 3440, 3450, 3465 (4 OH), 3387, 3395 (2 NH), 1728 in vitro primary antitumor screening on hepato-cellular carci-
(CO carboxylic acid), 1650, 1645, 1640 (3C]N) cmꢀ1. MS: m/z ¼ noma (HEPG2) and Caucasian breast adenocarcinoma (MCF7)
550 [M+ ꢀ 2]; anal. calcd for C29H24N6O6: C, 63.04; H, 4.34; N, (American Type Culture Collection). Cell viability was assessed
15.21; found: C, 63.09; H, 4.26; N, 15.25.
by the mitochondrial-dependent reduction of yellow MTT (3-
2-((2,8-Dihydroxy-4-acetyl-[1,2,4]triazolo[1,5-a]quinolin-9-yl)- (4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) to
methylamino)-3-(1H-benzo-4-yl)-9-(1H-benzimidazole-carboxylic purple formazan.36
acid) (18). Yield: 63%; m.p. 211–215 ꢁC. 1H-NMR (300 MHz,
Procedure. All the following procedures were performed in a
DMSO): d 2.26 (3H, s, CH3), 4.30 (2H, d, CH2–), 4.48 (2H, s, CH2–), sterile area using a laminar ow cabinet bio-safety class II level
5.70 (1H, t, CH–), 6.65–6.90 (3H, m, Ar-H), 7.10–7.25 (3H, m, Ar- (Baker, SG403INT, and Sanford, ME, USA). Cells were sus-
H), 7.35–7.60 (5H, m, Ar-H), 9.48 (1H, br s, NH–), 9.80 (1H, br s, pended in RPMI 1640 medium for HEPG2 and MCF7. The
NH-benzimidazole), 10.95 (1H, br s, COOH), 11.45 (1H, br s, OH- media are supplemented with 1% antibiotic–antimycotic
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25, 12.5, 6.25, 3.125, 0.78 and 1.56 mg mlꢀ1. Aer 48 h of incu-
bation, the medium was aspirated, 40 ml of MTT salt (2.5 mg
mlꢀ1) were added to each well and incubated for a further 4 h at
37 ꢁC under 5% CO2. To stop the reaction and dissolve the
formed crystals, 200 ml of 10% sodium dodecyl sulphate (SDS) in
deionized water was added to each well and incubated over-
night at 37 ꢁC. A positive control that is composed of 100 mg
mlꢀ1 was used as a known cytotoxic natural agent that gives
100% lethality under the same conditions.37
The absorbance was then measured using a microplate
multi-well reader (Bio-Rad Laboratories Inc., Model 3350,
Hercules, CA, USA) at 595 nm and a reference wavelength of 620
nm. Statistical signicance was tested between samples and a
negative control (cells with vehicle) using an independent t-test
by the SPSS 11 program. DMSO is the vehicle used for dissolu-
tion of plant extracts, and its nal concentration on the cells
was less than 0.2%. The percentage of change in viability was
calculated according to the formula:
Table 1 In vitro cytotoxic effect of the newly synthesized compounds
against hepatocellular carcinoma cell line (HEPG2) at 100 mg mlꢀ1 and
IC50 value of promising compounds
a
Compound
IC50 (mg mlꢀ1
)
% cytotoxicity at 100 mg mlꢀ1
4
5
6
7
—
—
—
13.6% at 100 mg mlꢀ1
29.3% at 100 mg ꢀm1lꢀ1
0% at 100 mg mlꢀ1
0% at 100 mg ml
—
8
14
15
16
17
19
—
—
—
54.5
—
62.5
—
—
0% at 100 mg mlꢀ1
40.2% at 100 mg mlꢀ1
0% at 100 mg mlꢀ1
87.5% at 100 mg mlꢀ1
33.3% at 100 mg mlꢀ1
89.2% at 100 mg mlꢀ1
1% at 100 mg mlꢀ1
0%
DMSO
Negative control
Doxorubicin
21.6
a
IC50 is dened as the concentration which results in a 50% decrease in
cell number as compared with that of the control structures in the
absence of an inhibitor.
((Reading of extract/reading of negative control) ꢀ 1) ꢂ 100.
Table 2 In vitro cytotoxic effect of the newly synthesized compounds
against breast adenocarcinoma cell line (MCF-7) at 100 mg mlꢀ1 and
IC50 value of promising compounds
A probate analysis was carried out for IC50 determination
using the SPSS 11 program.
The percent inhibition of cell viability was determined with
reference to the control values. The data were subjected to
linear regression analysis and the regression lines were plotted
for the best t. The IC50 (inhibition of cell viability) concentra-
tions were calculated using the respective regression equation.
From the results obtained in Tables 1 and 2, it was observed
that some of the tested compounds showed signicant activity
against HEPG2 and MCF-7 cancer cell lines, where some
compounds showed moderate to low activity.
The results of the antitumor screening allow for the
following assumptions about the structure–activity relationship
(SAR) of the synthesized compounds.
For the cytotoxic activity against HEPG2 (liver carcinoma cell
line): the results show that compounds 19 and 16 were almost
comparable with doxorubicin (Table 1), which may be corre-
lated with the presence of the imidazolyl moiety in both
compounds. For the cytotoxic activity against MCF-7 (cells from
breast cancer):
The cytotoxic activity of the synthesized tested compounds 4,
19, 8, 17 and 12 against MCF-7 ranged from potent to moderate
activity in a descending order, respectively. Compounds 4 and 8
are open chain substituted triazolo methylamino carboxylic
acids, whereas compound 19 comprises an imidazole as well as
a benzimidazole moiety.
Compounds 12 and 17 are both related to compound 19 as
they contain the benzoimidazole carboxylic acid, which might
be attributed to the cytotoxic activity.
Compound 7 is also related to compounds 4 and 8 being a
triazolo methylamino carboxylic acid, but the activity might be
less due to the absence of the open chain noticed in compounds
4 and 8.
Therefore, the appreciable cytotoxicity of the triazolo quin-
olines, especially compounds 19 and 16, against the HEPG2 cell
Compound
IC50a (mg mlꢀ1
)
% cytotoxicity at 100 mg ml-1
4
5
6
7
8
9
10
11
12
13
17
18
41.4
—
—
80.9
62.0
—
—
—
76.4
—
74.1
—
93.9% at 100 mg mlꢀ1
20.7% at 100 mg mlꢀ1
14.1% at 100 mg mlꢀ1
64.3% at 100 mg mlꢀ1
85.3% at 100 mg mlꢀ1
30.9% at 100 mg mlꢀ1
16.1% at 100 mg mlꢀ1
5.7% at 100 mg mlꢀ1
68.3% at 100 mg mlꢀ1
11.2% at 100 mg mlꢀ1
67.1% at 100 mg mlꢀ1
50.9% at 100 mg mlꢀ1
83.2% at 100 mg mlꢀ1
3% at 100 mg mlꢀ1
0%
19
DMSO
Negative control
Doxorubicin
52.1
—
—
26.1
a
IC50 is dened as the concentration which results in a 50% decrease in
cell number as compared with that of the control structures in the
absence of an inhibitor.
mixture (10 000 U mlꢀ1 potassium penicillin, 10 000 mg mlꢀ1
streptomycin sulfate and 25 mg mlꢀ1 amphotericin B), 1% L-
glutamine and 10% fetal bovine serum and maintained at 37 ꢁC
under 5% CO2.
Cells were batch cultured for 10 days, then seeded at a
concentration of 10 ꢂ 103 cells per well in fresh complete
growth medium in 96-well microtiter plastic plates at 37 ꢁC for
24 h under 5% CO2 using a water jacketed carbon dioxide
incubator (Sheldon, TC2323, Cornelius, OR, USA). Media was
aspirated, fresh medium (without serum) was added and cells
were incubated either alone (negative control) or with different
concentrations of sample to give nal concentrations of 100, 50,
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line may be attributed to the imidazole moieties as well as the
benzimidazole moiety.
As shown in Tables 3 and 4, the synthesized compounds
showed different antimicrobial and antifungal activities.
Table 3, representing results obtained for compounds 4–13,
We can also relate the cytotoxicity of compounds 4 and 8
against breast cancer cell line MCF-7 to the aliphatic moiety of shows that compounds 9 and 11 exhibited strong antibacterial
the amino carboxylic acid; the steric hindrance of the imidazole activities against Gram negative bacterium Escherichia coli
moiety in compound 7 may contribute to its weaker activity.
reaching a 20 mm and a 25 mm clear zone, respectively,
exceeding the activity of the reference antibiotic ampicillin (15
mm), which is a good result of medical value.
Table 4 shows the results obtained for compounds 14–19.
Compound 15 showed strong antibacterial activity against
Escherichia coli (Gram negative stain bacterium) with double the
value of the reference antibiotic used reaching a 30 mm clear
zone. Thus, we can report this novel compound as an excellent
formula against Gram negative highly pathogenic Escherichia
coli and stronger than compounds 9 and 11.
Compounds 18 and 19 exhibited good inhibitory activity
against Gram negative bacterium Escherichia coli. These results
are of great value for pharmaceutical and biological control
applications.
The outer membrane of Gram negative bacteria comprises a
complex liposaccharide (LPS) that acts as an endotoxin, protects
the bacteria from several antibiotics, dyes, and detergents and
provides these bacteria with resistance to lysozyme and peni-
cillin. However, alternative medicinal treatments such as lyso-
zyme with EDTA and the antibiotic ampicillin have been
developed to combat the protective outer membrane of some
pathogenic Gram-negative organisms. Other drugs can also be
used, the signicant ones being chloramphenicol, strepto-
mycin, and nalidixic acid (Fig. 1).
5. Antimicrobial activity
The antibacterial and antifungal activities of the synthesized
compounds were tested in vitro in comparison with ampicillin as
a reference drug using the standard agar disc diffusion method
against ve bacterial species and four fungal species. The anti-
bacterial and antifungal activities were carried out in the
Microbial Chemistry Department, National Research Center,
using the diffusion plate method. A lter paper sterilized disc
saturated with a measured quantity (25 ml) of each sample (1 mg
mlꢀ1 nal concentration) was placed on a plate (9 cm diameter)
containing a solid bacterial medium (nutrient agar) or a fungal
medium (potato dextrose agar) that had been seeded with the
spore suspension of the tested organism. Aer incubation at 37
ꢁC for 24 h for bacteria (in case of fungi, at 25 ꢁC for 72 h), the
diameter of the clear zone of inhibition surrounding the sample
was taken as a measure of the inhibitory power of the sample
against the particular tested organism (% inhibition ¼ sample
inhibition zone (mm)/plate diameter ꢂ 100).
All the tested compounds were solubilized in DMSO as a
solvent that has zero inhibition activity.38–41
Based on our results in this research paper we introduce
compounds 15, 11 and 9 as effective agents against the Gram
negative pathogen Escherichia coli and suggest applying phar-
macological drug tests on them.
Compound 13, on the other hand, exhibited high antibac-
terial activity against Gram positive bacterium Staphylococcus
aureus, which was almost equal to the activity of the reference
antibiotic ampicillin.
The antibacterial activities of the tested compounds were
examined with Gram positive bacteria, Bacillus cereus and
Staphylococcus aureus, and Gram negative bacteria Escherichia
coli, Pseudomonas aeruginosa and Salmonella typhimurium.
Antifungal activity was examined against Candida albicans,
Fusarium sp., Rhizoctonia sp., and Mucor sp.
The obtained results were compared with the reference
antibiotic ampicillin that was purchased from the Egyptian
market (Pzer Company, Cairo).
Compound 17 was the most active against Gram positive
bacterium Bacillus cereus; moreover, it exhibited a good inhib-
itory effect against Gram negative bacterium Escherichia coli.
The results revealed that the compounds showed varying
degrees of inhibition against the tested microorganisms.
Table 3 The antibacterial and antifungal activities of the tested compounds 4, 5, 6, 7, 8, 9, 10, 11, 12 and 13
Inhibition zone diameter (mm per mg sample)
Compound number
Gram stain
reaction
Reference antibiotic
ampicillin
Microorganism
4
5
6
7
8
9
10
11
12
13
Bacillus cereus
Escherichia coli
Pseudomonas aeruginosa
Staphylococcus aureus
Salmonella typhimurium
Fusarium sp.
Rhizoctonia sp.
Mucor sp.
Candida albicans
Positive
Negative
Negative
Positive
Negative
Fungus
Fungus
Fungus
Fungus
5
3
0
10
12
35
0
0
0
0
0
10
0
0
0
12
5
3
0
11
0
8
0
0
0
10
5
0
10
9
0
16
0
0
20
0
0
0
0
0
0
0
0
0
9
0
0
0
22
0
0
25
0
0
0
0
0
0
0
10
0
15
10
0
0
0
0
12
0
12
0
28
0
0
22
0
25
15
45
30
40
25
28
25
35
10
12
8
9
15
0
0
25
0
12
0
10
10
10
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Table 4 The antibacterial and antifungal activities of the tested compounds 14, 15, 16, 17, 18 and 19
Inhibition zone diameter (mm per mg sample)
Compound number
Gram stain
reaction
Reference antibiotic
ampicillin
Microorganism
14
15
16
17
18
19
Bacillus cereus
Escherichia coli
Pseudomonas aeruginosa
Staphylococcus aureus
Salmonella typhimurium
Fusarium sp.
Rhizoctonia sp.
Mucor sp.
Candida albicans
Positive
Negative
Negative
Positive
Negative
Fungus
Fungus
Fungus
Fungus
8
10
0
0
0
0
0
0
0
0
30
0
0
0
0
15
0
10
11
0
0
0
0
0
10
10
35
15
0
25
0
0
32
0
0
15
0
0
0
0
0
0
0
0
15
0
0
18
0
10
0
0
25
15
45
30
40
25
28
25
35
0
0
Fig. 1 Gram positive and Gram negative bacteria: a Gram positive bacterium has a thick layer of peptidoglycan (left). A Gram negative bacterium
has a thin peptidoglycan layer and an outer membrane (right) (http://www.digitalproteus.com).
Compound 17 also exhibited an antifungal activity against chemical and pharmacological properties. The present study
Rhizoctonia sp. This compound has the advantage of distinct reports an efficient and convenient synthesis of a new series of
effects on both tested bacterial and fungal pathogens.
selenadiazoles and triazolo[1,5-a]quinoline derivatives. The
Gram-positive bacteria lack an outer membrane but are synthesized compounds were screened in order to evidence
surrounded by layers of peptidoglycan many times thicker than anticancer and antimicrobial activities. Some of them such as
those found in Gram-negative bacteria (Fig. 1).
compound 4 exhibited strong inhibitory effects against breast
Special importance must be given to novel agents having the cancer (MCF-7); compounds 8 and 19 showed moderate
ability to inhibit the growth of such pathogens to overcome the signicant cytotoxic activities towards the tested cell lines, in
pathogens' resistance to normally used antibiotics.
particular with the liver carcinoma cell line HEPG2; and
Compound 4 showed superior antifungal activity (against compounds 16 and 19 gave moderate activity. Compounds 4–19
Fusarium sp.) that was achieved by a clear zone reaching 35 mm. were bio-assayed in vitro against four kinds of phytopathogenic
The same compound 4 displayed good activity against the fungi (Fusarium sp., Rhizoctonia sp., Mucor sp. and Candida
Candida albicans pathogen. This compound may affect an albicans) and ve Gram positive and Gram negative microor-
essential component of the fungal cell membrane and thus ganisms (Bacillus cereus, Escherichia coli, Pseudomonas aerugi-
inhibit its growth. The three major groups of antifungal agents nosa, Salmonella typhimurium and Staphylococcus aureus). The
in clinical use, i.e., azoles, polyenes, and allylamine/thio- results showed that some of the synthesized compounds
carbamates, all owe their antifungal activities to inhibition of exhibited strong antifungal activities, among which compounds
the synthesis of or direct interaction with ergosterol, which is 4 and 17 displayed more potent activities against Fusarium sp.
the predominant component of the fungal cell membrane and Rhizoctonia sp. Compounds 9 and 11 exhibited high activity
(Parks and Casey, 1996).42
against Escherichia coli, whereas compound 13 exhibited high
activity against Staphylococcus aureus. Compound 17 is the most
active compound against Bacillus cereus, Escherichia coli and
Rhizoctonia sp., and compound 15 gave the highest activity
against Escherichia coli, but compounds 18 and 19 also dis-
played high activity against Escherichia coli.
6. Conclusion
Quinoline and its derivatives have always attracted both
synthetic and biological chemists because of their diverse
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17 Z. Luo, C. Zeng, F. Wang, H. He, C. Wang, H. Du and L. Hu,
Chem. Res. Chin. Univ., 2009, 25(6), 841.
Conflict of interest
18 P. A. Winstanley, S. A. Ward and R. W. Snow, Microbes Infect.,
2002, 4, 157.
The author(s) conrms that this article content has no conicts
of interest.
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Acknowledgements
The authors are thankful to the National Research Center, Egypt
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