Constituents of Plantago major subsp. intermedia with antioxidant and anticholinesterase capacities

The methanol extract of Plantago major subsp. intermedia (Gilib.) Lange afforded 4 known compounds, namely isomartynoside (1), 10-hydroxymajoroside (2), beta-sitosterol (3), and ursolic acid (4). Their structures were established by spectroscopic methods. After determination of the total phenolic and flavonoid contents of the methanol extract, the antioxidant potentials of the crude extract and isolated compounds 1-4 were determined by beta-carotene bleaching, DPPH free radical and ABTS cation radical scavenging, and cupric reducing antioxidant capacity methods. The methanol extract, rich in phenolic contents, indicated the same DPPH free radical scavenging activity (72% inhibition) as a standard compound, butylated hydroxytoluene, at 100 mu g/mL. Isomartynoside (1), a phenylpropanoid glycoside, showed the best inhibition of lipid peroxidation, ABTS cation radical scavenging activity, and cupric reducing antioxidant capacity among the tested samples. The anticholinesterase effects of the methanol extract and isolated compounds 1-4 were established using the Ellman method. A triterpenic acid, ursolic acid (4), exhibited moderate acetyl- (54.01 +/- 0.82%) and butyryl-cholinesterase (68.74 +/- 0.36%) inhibitory activity at 200 mu g/mL. Isomartynoside (1) was isolated here for the first time from a Plantago species; the antioxidant and anticholinesterase activities of P. major subsp. intermedia and compounds 1-2 were not previously determined.