Antioxidant and antiproliferative activity analysis of phenolic compounds from leaf extracts of Ocimum gratissimum AND Rosmarinus officinalis

Abstract

Cancer remains a challenge in Africa despite the recent therapeutic advances. Due to the side effects associated with the use of clinical drugs, Lamiaceae plants such as Ocimum gratissimum and Rosmarinus officinalis have been used as anti-proliferatives against various cancer cell lines, and their leaves used as traditional medicine in conditions such as diabetes, cancer, diarrhea and typhoid. Even then, limited studies exist on the nature and variability of polyphenols in these plants and their efficacy as anti-proliferatives on prostate, colorectal and cervical cancer cell lines. The aim of this study was to isolate, characterize and determine the antioxidant and anti-proliferative properties of phenolic compounds of crude leaf extracts of O. gratissimum and R. officinalis. Specific objectives were; to determine the total phenolic content (TPC) and antioxidant(AO) activity of crude organic extracts, to evaluate the in vitro antiproliferative activity of crude organic extracts against human prostate (DU145), cervical (HeLa229) and colorectal (CT26) cancer cell lines, to isolate and characterize phenolic compounds in bioactive crude organic extracts, to evaluate in vitro antiproliferative activity of phenolic crude isolates against DU145, HeLa229 & CT26, and to evaluate the in vitro cytotoxic activity of phenolic crude compounds. Experimental design was used in the study. The plant leaf samples were obtained from their cultivated areas in Wakiso district, Uganda by judgemental sampling. Crude organic extracts were obtained by maceration method using n-hexane, dichloromethane, ethylacetate and methanol and their total phenolic content and antioxidant activity determined by Folin Ciocalteu and 2,2-diphenyl-1-picryl-hydrazyl- hydrate (DPPH) methods respectively. In vitro antiproliferative activity of seven different concentrations (1000 to 1.37 μg/ml) of crude organic extracts against cancer cell lines was evaluated by MTT assay. The Methanol extract showed the highest antiproliferative bioactivity. The extract was fractionated using n-hexane, dichloromethane, ethylacetate and methanol, in their order of increasing polarity. The fractions were further isolated into polyphenols using Solid phase extraction (SPE) and characterized by Liquid chromatography mass spectrometry (LC-MS/MS). The isolates antiproliferative activity against cancer cell lines and cytotoxicity activity on normal vero cells was evaluated by MTT method. Doxorubicin was used as the positive control drug in all bioassays. Crude methanol extracts yielded the highest Total phenolic content (R. officinalis: 476.80± 0.40 μg/ml; O. gratissimum: 401.00±6.47 μg/ml) and % inhibition of DPPH (R. officinalis: 69.76 ± 0.09%; O. gratissimum: 61.26 ± 0.09%), with significant differences between the two plants extracts (p˂0.05). FTIR spectra confirmed presence of phenolic groups, alcohols, aromatics, nitro groups as well as carbonyl groups which is the reason for the various medicinal properties of these plants. Gallic acid, Rutin, Catechin & Quercetin compounds were found to be common in both plants’ extracts. Procyanidin, Carboxystrictosinedine, Isoferullic acid, Psychotrin, hydroxyplorentin, cephalin, Isoquercetin, Diadzin and hyperin were reported for the first time in O. gratissimum while 8 compounds were reported for the first time in R. officinalis which include Procyanidin, hydroxyplorentin, cephalin, Isoquercetin, Latifoliamide, Diadzin, hyperin and emetine. Also, methanol crude extracts had the highest antiproliferative activity on all cancer cell lines, and whose minimum inhibitory concentration (μg/ml) that killed 50% of cells (IC 50 ) was DU145-147.378, CT26-301.992 and HeLa229-432.4745 for R. officinalis and DU145- 104.839, CT26-586.683 and HeLa229-359.914 for O. gratissimum. The results obtained show that all the extracts under investigation were not toxic to normal vero cells, compared to the positive control which was doxorubicin drug (6.36 ± 0.45 μg/ml) which is potentially very toxic. Therefore, results show selective action and potential use of these plants to generate lead compounds for use in developing drugs against prostate, colorectal and cervical cancers. Further antiproliferative activity studies in pure compounds of O. gratissimum & R. officinalis as well as vivo models are recommended.