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Abstract Gymnocarpos decandrus Forssk., family Caryophyllaceae is a well-known grazing wild plant growing in the middle east region and north Africa. Recently several medicinal activities; anti-inflammatory, analgesic and diuretic were reported for G. decandrus.Forssk. The current investigation includes identification, authentication, phytochemical investigation and biological evaluation of G. aerial parts. The botanical study and DNA fingerprinting were performed for complete characterization, documentation as well as differentiation of G. decandrus Forssk. Moreover identification through the chemical constituents via HPLC/PDA/MS/MS that allowed for detection of 19 metabolites. Flavonoids were the major class of secondary metabolites. Application of various chromatographic and spectral techniques resulted in isolation and identification of 5,2’ -Dihydroxy-7,8,6’- trimethoxyflavone (rivularin), vitexin, protocatechuic acid, quercetin, isorhamnetin and quercetin 3-O-Ý-D-glucopyranoside (isoquercetin) from G. decandrus Forssk. Moreover, criteria for standardization of G. decandrus Forssk. extract were established including determination of proximate composition and nutritive value as well as analysis of carbohydrate, lipids and amino acid components. The safety margin (LD50 and chronic toxicity), antidiabetic, Ü-Glucosidase inhibitory effect, Coxsackie B4 antiviral activity and cytotoxic effects of the plant ethanolic extract were assessed |