Phytochemical Analysis, In vitro Assessment of Antioxidant Properties and Cytotoxic Potential of Thymus capitatus Essential Oil

Abstract

Essential oils (EOs) of medicinal plants became a matter of interest in scientific research because of their potential of utilization for several therapeutic purposes, The EO of Thymus capitatus has been traditionally used in treatment of several conditions related to skin, muscles, and respiratory system, to reduce fever, and to stimulate menstrual flow in addition to its anti-bacterial properties. To chemically analyze the components of the EO of Thymus capitatus and to evaluate its anti-cancerous and anti-oxidant activity on breast cancer cells (T47D), colorectal adenocarcinoma cells (CACO2), and normal human fibroblasts (MRC5). The EOs were extracted by hydro-distillation. Gas chromatography and mass spectrometry were used to analyze the extracts. Cell viability was assessed using three different assays; MTT, trypan blue and neutral red assays. Antioxidant activity was assessed using DPPH scavenging assay. Cell apoptosis was assessed using DNA fragmentation tests and Annexin V-propidium iodide assay. Morphological analysis of cells was assessed using light microscopy and DAPI staining. Twenty-two different compounds were identified in the essential oil of T. capitatus leaves including Thymol (44.34%) and Carvacrol (38.89%) in addition to 20 minor components. The oil extract exhibited a significant inhibitory action on the three cell lines. The viability assays showed significantly higher (p<0.05) IC50 in the fibroblast cell line than the other two cell lines. The selectivity index was higher in T47D cell line compared to CACO II cell line. For apoptosis assessment, DNA fragmentation in T47D cells was significantly correlated with the extract concentration. The membrane polarity was disrupted in an increasing manner (15.60±1.13, 35.73±1.69 and 46.53±2.51) as the extract concentration (20.22, 40.45, and 80.9µg/ml, respectively) increased. On the cellular level, morphological changes including cell shrinkage, chromatin condensation and nuclear fragmentation in addition to marginalization of chromatin, vacuolization of cytoplasm and formation of micronuclei was clearly visible. The extract showed high anti-oxidant activity with an IC50 of 6.4µg/ml. Anti-oxidant activity was dose-dependent; double IC50 (80.9µg/ml) exhibited the highest antioxidant activity whereas the lowest antioxidant activity was observed for the ½ IC50 (20.225µg/ml). The oil extract exerted an inhibitory action lipid peroxidation. Thymus capitatus EO is a potential anti-oxidant and anti-neoplastic agent against breast and colon carcinomas due to its high selectivity for cancer cells compared to normal cell lines.