METHODS: Samples of leaves, stems, flowers and roots from E. hirta were tested for total phenolic content, and flavonoids content and in vitro antioxidant activity by diphenyl-1-picrylhydrazyl (DPPH) assay and reducing power was measured using cyanoferrate method.
RESULTS: The leaves extract exhibited a maximum DPPH scavenging activity of (72.96±0.78)% followed by the flowers, roots and stems whose scavenging activities were (52.45±0.66)%, (48.59±0.97)%, and (44.42±0.94)%, respectively. The standard butylated hydroxytoluene (BHT) was (75.13±0.75)%. The IC(50) for leaves, flowers, roots, stems and BHT were 0.803, 0.972, 0.989, 1.358 and 0.794 mg/mL, respectively. The reducing power of the leaves extract was comparable with that of ascorbic acid and found to be dose dependent. Leaves extract had the highest total phenolic content [(206.17±1.95) mg GAE/g], followed by flowers, roots and stems extracts which were (117.08±3.10) mg GAE/g, (83.15±1.19) mg GAE/g, and (65.70±1.72) mg GAE/g, respectively. On the other hand, total flavonoids content also from leave had the highest value [(37.970±0.003) mg CEQ/g], followed by flowers, roots and stems extracts which were (35.200±0.002) mg CEQ/g, (24.350±0.006) mg CEQ/g, and (24.120±0.004) mg CEQ/g, respectively. HPTLC bioautography analysis of phenolic and antioxidant substance revealed phenolic compounds. Phytochemical screening of E. hirta leaf extract revealed the presence of reducing sugars, terpenoids, alkaloids, steroids, tannins, flavanoids and phenolic compounds.
CONCLUSIONS: These results suggeste that E. hirta have strong antioxidant potential. Further study is necessary for isolation and characterization of the active antioxidant agents, which can be used to treat various oxidative stress-related diseases.
AIM OF THE STUDY: The purpose of this study was to determine the in situ cytotoxicity effect P. macrocarpa fruit ethyl acetate fraction (PMEAF) and the underlying molecular mechanism of cell death.
MATERIALS AND METHODS: MDA-MB-231 cells were incubated with PMEAF for 24h. Cell cycle and viability were examined using flow cytometry analysis. Apoptosis was determined using the Annexin V assay and also by fluorescence microscopy. Apoptosis protein profiling was detected by RayBio® Human Apoptosis Array.
RESULTS: The AO/PI staining and flow cytometric analysis of MDA-MB-231 cells treated with PMEAF were showed apoptotic cell death. The cell cycle analysis by flow cytometry analysis revealed that the accumulation of PMEAF treated MDA-MB-231 cells in G0/G1 and G2/M-phase of the cell cycle. Moreover, the PMEAF exert cytotoxicity by increased the ROS production in MDA-MB-231 cells consistently stimulated the loss of mitochondrial membrane potential (∆Ψm) and induced apoptosis cell death by activation of numerous signalling proteins. The results from apoptosis protein profiling array evidenced that PMEAF stimulated the expression of 9 pro-apoptotic proteins (Bax, Bid, caspase 3, caspase 8, cytochrome c, p21, p27, p53 and SMAC) and suppressed the 4 anti-apoptotic proteins (Bcl-2, Bcl-w, XIAP and survivin) in MDA-MB-231 cells.
CONCLUSION: The results indicated that PMEAF treatment induced apoptosis in MDA-MB-231 cells through intrinsic mitochondrial related pathway with the participation of pro and anti-apoptotic proteins, caspases, G0/G1 and G2/M-phases cell cycle arrest by p53-mediated mechanism.