In Vitro Evaluation of Antioxidant, Anticancer, and Anti-Inflammatory Activities of Ethanolic Leaf Extract of Adenium obesum

Abstract

Adenium obesum commonly known as "desert rose" belongs to the family Apopcynaceae and has previously been reported for its anti-influenza, antimicrobial, and cytotoxic efficacies and well-known for their ethno-medicinal applications. In the present study, ethanolic extracts of A. obesum (AOE) were analyzed by gas chromatography-mass spectrometry (GC-MS) to identify the important phytochemical compounds. The GC-MS analysis of AOE detected the presence of 26 phytochemical compounds. This plant is traditionally used for the treatment of various diseases. In this report, the antioxidant, anti-inflammatory, and anticancer activities of ethanolic leaf extract from A. obesum (AOE) were studied. The antioxidant potential of ethanolic extract of AOE was examined by different antioxidant assays, such as antioxidant capacity by the DPPH, ABTS, superoxide, hydroxyl radical scavenging, and lipid peroxidation inhibition assays. The antioxidant activities of various reaction mixtures of AOE were compared with a reference or standard antioxidant (ascorbic acid). In addition, we also evaluated the anticancer activity of AOE, and it was observed that AOE was found to be cytotoxic against A549 lung cancer cells. It was found that AOE inhibited the viability of A549 lung cancer cells by inducing nuclear condensation and fragmentation. Furthermore, ethanolic AOE demonstrated the anti-inflammatory potential of AOE in murine alveolar macrophages (J774A.1) as an in vitro model system. AOE showed its potential in reducing the levels of inflammatory mediators including the proinflammatory cytokines and TNF-α. The results obtained in the present investigation established the antioxidant, anticancer, and anti-inflammatory potency of AOE, which may account for subsequent studies in the formulation of herbal-based medicine.

Keywords: TNF-α; anti-inflammatory; anticancer; antioxidant; cytokines.

FIGURE 1

GC-MS chromatogram of the ethanolic extract of Adenium obesum leaves.

FIGURE 2

Effect of AOE on the growth of A549 lung cancer cells. (A) Percent (%) cell viability of A549 cells after AOE treatment at concentrations of 100–400 μg/ml after 24 h. (B) Phase-contrast images of AOE-treated A549 cells for 24 h. Data communicated constitutes the mean ± SEM of individual experiments performed thrice in triplicate. The level of significance among different AOE-treated groups was determined using one-way ANOVA and Dunnett’s post-hoc in comparison with the untreated control where *represents p<0.05, ***p<0.001; Scale bar = 100 μm, and magnification = 20X

FIGURE 3

(A) AOE-mediated induction of apoptosis within treated human lung cancer A549 cells. Increased levels of nuclear condensation and apoptosis are depicted within photomicrographs of vehicle control and AOE-treated (100–400 µg/ml) A549 cells stained with Hoechst 33342 by red arrows. (B) Enhanced intracellular ROS generation in DCHF-DA-stained A549 cells treated with AOE for 24 h. Scale bar = 100 μm and magnification = 20X

FIGURE 4

AOE treatment-induced consequences on activation of caspase-9 and -3 within A549 cells (A) percent (%) activation of caspase-9 and -3 activities post AOE exposure at 100–400 µg/ml concentrations, (B-E) mRNA expression levels of pro- and anti-apoptotic genes within AOE-treated A549 cells. (E) Decrease in NIR fluorescence in AOE-treated cells after staining with Mito-NIR dye, suggesting depolarization of ΔΨ_m.

FIGURE 5

Potential of AOE in deflating the LPS-mediated inflammatory response as characterized by the levels of (A) IL-1β, (B) TNF-α, (C) IL-6, and (D) PGE2 within J774A.1. . Data communicated constitute the mean ± SEM of individual experiments performed thrice in triplicates. The level of significance among different AOE-treated groups was determined using one-way ANOVA and Dunnett’s post-hoc in comparison with untreated control where *represents p<0.05 and **p<0.01