Radioprotection by Pilea microphylla (L.): a free radical mechanistic and molecular approach

Abstract

The present work involved isolation, identification and characterization of compounds present in Pilea microphylla (PM) followed by assessment of their free radical scavenging and radioprotective ability. Six phenolic compounds namely, quercetin-3-O-rutinoside (PMC-1), 3-O-caffeoylquinic acid (PMC-2), luteolin-7-O-glucoside (PMC-3), apigenin-7-O-rutinoside (PMC-4), apigenin-7- O-β-D-glucopyranoside (PMC-5) and quercetin (PMC-6) were isolated. Compounds were characterized by 1D, 2D NMR techniques and high-resolution LC-MS. PMC-1, PMC-2, PMC-3 and PMC-6 exhibited significant antioxidant potential in scavenging free radicals and prevented lipid peroxidation (LPO). PMC-1, PMC-2, PMC-3 and PMC-6 attenuated levels of reactive oxygen species, LPO, cytotoxicity and DNA damage, in irradiated V79 cells. Inhibition of LPO is presumably due to 3´,4´-catechol ortho-dihydroxy moiety in the B-ring, which has a strong affinity for phospholipid membranes. The reaction of radiolytically generated hydroxyl radical with protein, lipid or DNA molecules was also prevented by PMC-1. PMC-1 pretreatment significantly decreased the apoptotic markers in irradiated V79 cells. PMC-1, at 25 mg/kg/i.p., 30 min prior to γ- radiation, prevented the depletion of endogenous antioxidants in mice, protected gastrointestinal tract and hematopoietic system, thus improving overall survival. PMC-1 attenuated the radiation-induced apoptosis in spleenocytes and prevented DNA damage in peripheral blood leukocytes. PMC-1 also reversed the changes by increasing ERK and decreasing BAX expressions compared to radiation control. The cytoprotective and antigenotoxic activity of the active fraction of PM (PM1) was compared with its active polyphenolic constituents viz., quercetin-3-Orutinoside, quercetin and 3-O-caffeoylquinic acid (equivalent to quantities present in PM1-25 µg/ml). The combination of polyphenols protected irradiated V79 cells more effectively than individual constituents. To conclude, radioprotection is possibly a synergistic effect of the phytochemicals present in the herbal extract, rather than any single component.