Abstract
This report highlights the phytochemical analysis, antioxidant potential and anticancer activity against breast carcinoma of 70% methanolic extract of lichen, Parmotrema reticulatum (PRME). Phytochemical analysis of PRME confirms the presence of various phytoconstituents like alkaloids, carbohydrates, flavonoids, glycosides, phenols, saponins, tannins, anthraquinones, and ascorbic acid; among which alkaloids, phenols and flavonoids are found in abundant amount. High performance liquid chromatography (HPLC) analysis of PRME revealed the presence of catechin, purpurin, tannic acid and reserpine. Antioxidant activity was evaluated by nine separate methods. PRME showed excellent hydroxyl and hypochlorous radical scavenging as well as moderate DPPH, superoxide, singlet oxygen, nitric oxide and peroxynitrite scavenging activity. Cytotoxicity of PRME was tested against breast carcinoma (MCF-7), lung carcinoma (A549) and normal lung fibroblast (WI-38) using WST-1 method. PRME was found cytotoxic against MCF-7 cells with an IC50 value 130.03±3.11 µg/ml while negligible cytotoxicity was observed on A549 and WI-38 cells. Further flow cytometric study showed that PRME halted the MCF-7 cells in S and G2/M phases and induces apoptosis in dose as well as time dependent manner. Cell cycle arrest was associated with downregulation of cyclin B1, Cdk-2 and Cdc25C as well as slight decrease in the expression of Cdk-1 and cyclin A1 with subsequent upregulation of p53 and p21. Moreover PRME induced Bax and inhibited Bcl-2 expression, which results in increasing Bax/Bcl-2 ratio and activation of caspase cascade. This ultimately leads to PARP degradation and induces apoptosis in MCF-7 cells. It can be hypothesised from the current study that the antioxidant and anticancer potential of the PRME may reside in the phytoconstitutents present in it and therefore, PRME may be used as a possible source of natural antioxidant that may be developed to an anticancer agent.
Highlights
The free radicals are generated in various biological systems and in the human body in the form of reactive oxygen species (ROS) and reactive nitrogen species (RNS)
Potassium persulfate (K2S2O8), 2-deoxy-2ribose, ethylene diammine tetraacetic acid (EDTA), ascorbic acid, trichloroacetic acid (TCA), mannitol, nitro blue tetrazolium (NBT), reduced nicotinamide adenine dinucleotide (NADH), phenazine metho-sulfate (PMS), sodium nitroprusside (SNP), 1,10-phenanthroline, sulphanilamide, naphthyl ethylenediamine dihydrochloride (NED), L-histidine, lipoic acid, sodium pyruvate, quercetin and ferrozine were obtained from Sisco Research Laboratories Pvt
High performance liquid chromatography (HPLC) grade acetonitrile, ammonium acetate, hydrogen peroxide, potassium hexacyanoferate, Folin-ciocalteu reagent, sodium carbonate, mercuric chloride, potassium iodide, anthrone, vanillin, thiourea, 2,4-dinitrophenylhydrazine, sodium hypochlorite, aluminium chloride, xylenol orange, butylated hydroxytoluene (BHT), N,N- dimethyl-4nitrosoaniline and BCIP/NBT substrate were taken from Merck, Mumbai, India. 1,1-diphenyl-2-picrylhydrazyl (DPPH), gallic acid, (+) catechin, curcumin, RNAase A, 49,69-diamidino-2 phenylindole and Triton X-100 were obtained from MP Biomedicals, France
Summary
The free radicals are generated in various biological systems and in the human body in the form of reactive oxygen species (ROS) and reactive nitrogen species (RNS). Antioxidants, possible protective agents, can be considered to ease from oxidative damage caused by free radicals in the human body and retarding the progress of many chronic diseases including ageing and cancer [2,3,4]. These natural antioxidants could modify the behaviour of cancer cells by altering their redox environment [5,6] as well as reduce their genetic instability and may be considered useful in cancer treatment [7]. Major attention is being given to search for better and safer antioxidants of natural origin, which may raise the efficiency of cancer treatment
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