Antioxidant and Cytotoxicity Potential of Six Synthesized Chalcones

Background: Chalcones are open-chain flavonoids which display a large number of pharmacological activities such as cytotoxic, anti-inflammatory including antioxidant. The objective of this study was to assess antioxidant and cytotoxic activity of six synthesized chalcones. Methodology: For the current experiments, 1,3-diphenylpropenone (compound R) was used as molecular model to synthetize six compounds, namely three benzyl-benzimidazolyl-chalcones (U1, U2, WAC1) and three imidazopyridinyl-chalcones (V1, V2, V3). All the compounds were evaluated for their ability to scavenge the stable free ABTS radical cation, according to the method develop by Choong et al. In addition, the cytotoxicity test described by Price et al., was performed using healthy human cell line, then in human malignant cell lines (HEP-2, A549). Results: All synthesized chalcones reduced the ABTS radical cation. Indeed, benzyl benzimidazolyl compounds WAC1, U1, U2, by developing respectively 39.61%, 66.09%, and 84.20% percentages of reduction, showed an antioxidant effect 6, 11 and 14 times greater than the compound R (6.14%). As a result, imidazopyridinyl-chalcones compounds, namely V1, V2 and V3 reduced the ABTS radical cation at 91.62%, 99.84% and 97.45% respectively, being 15 and 16 times more active than the compound R. About cytotoxicity, V2 inhibited not significantly HEP-2 malignant cells growth at 48.64%, compared to the standard product, i.e. doxorubicin that inhibited the growth of the same cells at 42.37%. WAC1 inhibited significantly the growth of A549 malignant cells at 89.53%, more than doxorubicin which percentage of growth inhibition was 71.58%. Conclusion: The presence of the α, β-unsaturated carbonyl system (or 1,3-diphenylpropenone) along with a benzimidazole or imidazopyridine heterocyclic ring is likely to contribute to both cytotoxic and antioxidant activities of these compounds. How to cite this paper: Kouakou, S.L., Ouattara, M., N’Guessan, J.P., Coulibaly, S., Irié-N’Guessan, A.G. and Kouakou-Siransy, G. (2018) Antioxidant and Cytotoxicity Potential of Six Synthesized Chalcones. Pharmacology & Pharmacy, 9, 536-546. https://doi.org/10.4236/pp.2018.912042 Received: November 1, 2018 Accepted: December 23, 2018 Published: December 26, 2018 Copyright © 2018 by authors and Scientific Research Publishing Inc. This work is licensed under the Creative Commons Attribution International License (CC BY 4.0). http://creativecommons.org/licenses/by/4.0/ Open Access S. L. Kouakou et al. DOI: 10.4236/pp.2018.912042 537 Pharmacology & Pharmacy


Introduction
Chalcones are widely present in nature as important intermediates in the flavonoid biosynthetic pathway in various ferns and plant species [1]. The basic structure of chalcones contains a C6-C3-C6 backbone including two aromatic rings linked by an α, β-unsaturated carbonyl group [1].
Prompted by all these considerations, in an effort to design a unique template endowed both with radical scavenger properties and cytotoxicity activities, we replaced a phenyl ring of chalcone with heterocyclic rings namely imidazopyridine and benzimidazole. The purpose of the herein study was to evaluate the antioxidant and cytotoxic activity of six synthesized chalcones in the perspective to propose them as forward models on which we can rely for the development of new molecules effective against cancers.

Samples under Study
The six compounds tested, i.e. benzimidazolyl-chalcones (V1, V2, WAC1), and imidazopyridinyl-chalcones (V1, V2, V3), including molecular model (compound R), were synthesized and provided as pure powder by the Department of Organic and Therapeutic Chemistry of Training and Research Unit of Pharmaceutical and Biological Sciences (Côte d'Ivoire). From the molecular model (1,3-diphenylpropenone or compound R: Figure 1), chemical modifications were made on the aryl groups (ring A and B) to obtain the compounds under study.
The ring A replaced by benzimidazole or imidazopyridine rings, different substituted in ring B with electro donor element, allowed the synthesis of benzimidazolyl-chalcone derivatives (U1, U2, WAC1) and imidazopyridinyl-chalcones (V1, V2, V3). All of the chalcone hybrids were characterized by nuclear magnetic resonance (300 MHz for 1H and 75 MHz for 13C) on a Brucker Avance 300. Mass spectra were recorded on a JEOL JMS DX300 spectrometer on mode ESI, and details on their synthesis and structural information are located in the supporting information [8] [9]. These products were in powder of different colors, insoluble in water and the usual organic solvents, except for dimethyl sulfoxide (DMSO).

Antioxidant Ability Test
We selected the decolourisation assay of ABTS •+ cationic radical for antioxidant potential evaluation, because it is suitable for the screening of hydrophilic as well as lipophilic molecules [10] [11].

ABTS Test
The test was performed according to Choong et al. [12]. ABTS [2,2'-azino-bis (3-ethylbenzothiazolin-6-sulfonic acid)] solution produces the ABTS •+ radical cation with a green blue colour in solution, for which the absorbance is measured at 730 nm. Molecules that have an antioxidant ability, by donating an electron, reduce the ABTS •+ cation and turn the green blue reaction medium to a completely discoloured solution. The amount of antioxidant in the sample test is therefore inversely proportional to the formation of the ABTS.

Obtention of Radical Cation ABTS •+
The ABTS •+ radical cation is obtained by mixing volume-by-volume (1:1 v/v) of a stock solution ABTS 7.0 mM with 2.6 mM potassium persulfate solution, followed by incubation in the dark for 12 to 16 hours. The absorbance measured by spectrophotometry was adjusted between 1.0 and 1.5 at 734 nm, adding 60 mL of phosphate buffer solution (pH 7.4) to 1 mL of solution containing the radical ABTS •+ .

Procedure of the ABTS Method
The tests were carried out according to the following steps:

Cytotoxic Capacity
It was carried out at Chatenay Malabry biology laboratory (Paris-France). The evaluation focused on a healthy umbilical endothelial cell line (HUVEC) and two (2) malignant lines of human origin, namely a liver cancer line (HEP-G2), and a lung cancer line (A549).

Cell Culture and Cytotoxicity Tests
HUVEC, HEP-G2, A549 cells were cultured in specific media supplemented with 10% Foetal Calf Serum (FCS) and 1% Penicillin Streptomycin (PS). The autotoxicity test with MTT was carried out according to the method described by Price et al. [13].

Procedure of the Cytotoxicity Tests
The tests were carried out according to the following steps: The results, expressed as percentage of inhibition cells growing, were calculated at using the same formula of antioxidant percentage inhibition. The codification for statistically significant difference was: *: 0.01 < p ≤ 0.05; **: 0.001 ≤ p ≤ 0.01; ***: p < 0.001.

Discussion
Antioxidant screening showed that in benzimidazolyl-chalcones series, the introduction of a phenyl group on pyrrolic-N (U1), or a hydroxyl group substitution in position 2 (U2) of N-benzylbenzimidazolyl-chalcones, allowed these compounds scavenging the ABTS •+ with a potent respectively 11 and 14 times greater than that of the molecular model. Our results are compatible with those reported by Murti et al. [14], who specified in their study that the antioxidant activity of benzimidazolyl-chalcones was around 39% to 70%. Also, our results suggest that the presence of electron-donating substituents type hydroxyl on benzylbenzimidazolyl-chalcone (U2), is favorable for the enhancement of antioxidant potency, as revealed by the work of Prakash et al. [15]; Ghawalkar et al. [16]; and Lembege et al. [17].
In imidazopyridinyl-chalcones series, the introduction of hydroxyl group at The mechanism action suggested for V2, whose cytotoxic effect was similar to that of doxorubicin on HEP-G2, is that it interacts as DNA intercalating agents leading to cell death [18].
The same mechanism action could also be attributed to WAC1, but the fact that its effect was superior to that doxorubicin on A549 cells, would allow us, to expand its action as an agent that can interact on the signalling pathways of cell transduction. Indeed, WAC1, would behave as an agonist of tumor necrosis factor α (TNFα) which selectively binds to TNFR1 receptors, activates the transduction of signals mediated by the apoptotic pathways TRADD and FADD, resulting in the stimulation of caspases which cleaves multiple proteins responsible for cell death [19] [20].
Our results are consistent with a recent work led by Kamal et al. [21], who showed that derivatives of chalcones vectorized by various heterocyclic rings like imidazopyridine, imidazopyrimidine or benzimidazole, exhibited significant antiproliferative activity against the A549 human lung malignant line.

Conclusions
The imidazopyridinyl-chalcones and benzimidazolyl-chalcones could be good profiles of antioxidant activity. Depending on the position of the phenolic group in phenyl ring, the radical scavenging activity of the imidazopyridinyl-chalcones decreased in the following order: 3-OH > 4-N(CH 3 ) 2 > 2-OH. In the benzimidazolyl-chalcones series, the presence of a 2-OH group on the phenyl group was favourable to antioxidant activities when another phenyl group was inserted on pyrrolic-N. However, insertion of a 5-chloro group in benzimidazole didn't enhance the activity of corresponding chalcone (WAC 1).
In addition, the best cytotoxic activity was provided by benzimidazolyl-chalcone bearing a halogen at the 5-position of the heterocycle (WAC1