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ABSTRACT:Bioactivit y-guided fracti onati on was perf ormed to eval uate t he anti oxidant ac tivit y of S tevi a rebaudiana B ert oni. T he met hanolic cr ude (MC) extract of leav es was fr acti onat ed using a m odified K upchan partiti oning m ethod. The antioxidant ac tivit y w as m easur ed f or t he MC and frac tions. Free radic al scav enging using 2, 2-Diphenyl-1-picryl-hydraz yl (D PPH) and tot al anti oxidant ac tivit y using phosphomol ybdenum c omplex m et hods wer e selec ted f or determinati on of t he anti oxidant activit y usi ng as corbic acid (A A) as a refer enc e. Preliminar y anti oxidant ac tivit y meas urem ents r ev eal ed t hat the dichloromethane frac tion w as of i nt erest . Cons equentl y, it was s ubject ed t o chrom at ographic s eparation and purification tec hniques. One m ajor pure c ompound w as isolat ed and its str uct ure elucid ation using NMR and HR MS c onfirmed it to b e 3' ,5, 7-trihydroxy-3, 4', 6-trimet hoxyflavone (cent aureidin). The IC 50 v alues of MC , butanol, m et hanol and dichlorom et hane fractions and c entaureidin (CT) wer e 120.6 ± 0. 58, 66. 5 ± 0. 49, 79. 5 ± 0. 44, 37.1 ± 0. 23, and 18. 4 ± 0. 44 µg/mL, r espectiv el y. The results wer e c ompar ed t o t hose of s ynt hetic antioxidants, but ylat ed hydroxyanisol e ( BH A) and but ylated hydroxyt oluene (BHT) wit h IC 50 values of 13. 7 ± 0. 09 and 33. 1 ± 1. 05 µg/mL, respectiv el y; w hile t hat of A A w as 6. 5 ± 0.25 µg/mL. T he hig hes t t ot al antioxidant ac tivit y eval uated b y t he p hosp hom ol ybdenum complex method was exhibited by t he purified compound (CT) with a v alue of 5028. 6 ± 645. 87 mg A A equiv alent/g sample. The stud y rev eals that c omponents of t his plant could b e a p otenti al alter native t o s ynt hetic antioxidants b y virtue of their anti oxidant properties and natur al origin. antioxidant activi t y; Ast erac eae; Stevia rebaudiana ; s ynthetic anti oxidant; nat ural antioxidant; Cent aur eidin.

Stevia rebaudiana Bertoni is an important medicinal plant; plant leaves are used as a source of non-caloric sweetener-stevioside, which is estimated to be 300 times sweeter than cane sugar. Seed germination of stevia is very low, so micropropagation is an essential prerequisite for stevia mass production and modification. Determined plant tissue culture conditions were applied to establish shoot cloning and synthetic seeds (synseeds) formation. Efficient shoot multiplication was established on MS medium with 0.25 mg/l BAP. While the best shoot formation frequency was obtained when nodal or shoot cuttings were sub cultured for the fifth time, addition of 1.7 mg/l of AgNO3 increased the values of shoot formation frequency and reduced verification. For production of artificial seeds, small nodal cuttings, about 3 mm long each, were coated using sodium alginate (4%) and CaCl2 (75 mM) solution as a gel matrix and a complexing agent, respectively. Shoot cuttings resulting from the conversion of synseeds or shoot culture were used to establish root formation on MS medium containing 1 mg/l IBA. Plantlets with extensive root systems were hardened and successfully established in the soil.