Study the effect of heavy metals and Nano TiO2 on stevioside and rebadioside A production in Stevia rebaudiana Bertoni hairy roots

Stevia rebaudiana Bert. is a medicinal plant with anti-oxidant, antimicrobial, antifungal and antiviral drug properties which contains two valuable secondary metabolites, stevioside and Rebadioside A, with very high sweetness capacity. In this research hairy roots were produced from S. rebaudiana leaf disks using two strains of 15834 and A4 A. rhizogenes. Then, the best hairy root line was selected and propagated. The effect titanium dioxide nanoparticles at 20, 40 and 60 mg/l after 0, 24, 48 and 72 h of elicitation and cobalt chloride and cadmium chloride each at 100 and 200 μmol after 2 and 8 h of elicitation were studied on the production and accumulation of stevioside and Rebadioside A. The results demonstrated that most treatments of elicitors at different times had significant effects on the production and accumulation of target metabolites, but titanium dioxide nanoparticles caused the highest production and accumulation of stevioside (96.88 mg/l and 24.28 mg/g DW), respectively at 40 mg/l after 24 h. However, at 60 mg/l and 48 h it had the highest effect on production and accumulation of Rebadioside A after 48 h (89.39 mg/l and 21.28 mg/g DW), respectively. Also the results demonstrated that treatment of cobalt chloride and cadmium chloride both at 100 μmol after 8 h had a strong effect on the production (114.19 and 183.98 mg/l) and accumulation (29.54 and 42.87 mg/g DW ) of stevioside, respectively. Cadmium chloride at 200 μmol had the highest effect on the production and accumulation of Rebadioside A after 2 h (105.51 mg/l and 25.42 mg/g DW), respectively. Meanwhile, no production of Rebadioside A was observed at 100 and 200 μmol in 8 h treatment.