In-vitro Cytotoxicity Evaluation of DL-limonene on Vero E6 cell line

Abstract

Studies have shown the beneficial effects of plant-derived small organic compounds, known as secondary metabolites, many of which have significant therapeutic potentials on cardiovascular disease, diabetes and certain types of cancer. A wide variety of active phytochemicals have been found to influence cellular functions, membrane permeability, and viral replication. Thus, naturally based pharmacotherapy such as DL-limonene may be a proper alternative for treating viral diseases such as Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV- 2) infection. However, there is a need for proper documentation of the toxicity and safety profile of these natural compounds. The aim of this study is to evaluate the cytotoxicity of DL-limonene on normal kidney cells of African Green Monkey (Vero E6 cell line),which is a non-cancerous cell line. In vitro cytotoxicity of DL-limonene on Vero E6 cells was evaluated by WST-8 assay. Afterwards, microscopic examination of the morphological alteration of the cells exposed to 5000.00 μM to 4.9 μM concentrations of DL-limonene was performed. The viability of cells treated with high concentrations (1250-5000 μM) of DL-limonene was reduced by more than 80% while increased cell growth and proliferation were observed in cells treated with lower concentrations (625-4.9 μM). Morphological changes, such as shrinking, blebbing, and cell rounding, were observed at varying concentrations (1250-5000 μM) of DL-limonene. In contrast, lower concentrations (625-4.9 μM) of DL-limonene showed no observable changes in the morphology of the cells. Therefore, DL-limonene is non-toxic to normal cells at low concentrations, supporting its safe utilisation for therapeutic applications.