Effect of Growth Regulators on Stevia rebaudiana Bertoni Callus Genesis and Influence of Auxin and Proline to Steviol Glycosides, Phenols, Flavonoids Accumulation, and Antioxidant Activity In Vitro

Abstract

Stevia is a plant containing many active compounds, but usually propagated by stem cuttings because of low seed-yield-germination ability. The aim of this study was to investigate the impact of plant-growth regulators on stevia callus induction and growth from somatic tissue, as well as to determine the effect α-naphthalene acetic acid (NAA) and proline (PRO) on the amount of stevioside, rebaudioside A, phenols, flavonoids, and antioxidant activity. Stem and leaf segments were inoculated on a Murashige and Skoog (MS) medium supplemented with different concentrations of NAA and 6-benzylaminopurine (BAP) for callus genesis. The amount of steviol glycosides (SGs) was evaluated using high-performance liquid chromatography (HPLC), and the amounts of total phenols, flavonoids, and antioxidant activity by spectrophotometric methods. The highest callus-induction frequency and callus-mass increase were obtained from the leaf explants in MS medium supplemented with 2.0 μM NAA. The highest amount of SGs, phenols, and flavonoids, and stronger antioxidant activity were determined in the cellular compounds of callus from leaf explant. PRO reduced the amount of SGs and flavonoids. The significantly highest amount of total phenolic compounds was obtained in the callus from leaf explants in the medium supplemented with 2.0 µM NAA and 2.0 µM PRO.

Callus Induction, Proliferation, Enhanced Secondary Metabolites Production and Antioxidants Activity of Salvia moorcroftiana L. as Influenced by Combinations of Auxin, Cytokinin and Melatonin

“…The present results are in close conformity with Rehman coauthors [38] who reported that callus biomass was significantly increased with the application of 2,4-D furthermore the addition of BAP enhanced the role of 2,4-D which resulted in increased biomass of Caralluma tuberculata callus. The results are supported by Blinstrubiene and coauthors [39] who observed that media when augmented with BAP and 2,4-D significantly increased the callus fresh weight of Stevia rebaudiana. These results are in agreement with Aghaei and coauthors [40] on Pistacia atlantica, Wang and Bao [41] on Viola wittrockiana and Manisha and Rajesh [42] on Tecomella undulata.…”

The Potential of Cold Plasma and Electromagnetic Field as Stimulators of Natural Sweeteners Biosynthesis in Stevia rebaudiana Bertoni

“…Steviol glycosides rebaudioside A (RebA) and stevioside (Stev) were separated and quantified using high-performance liquid chromatography (HPLC) [ 62 ]. An Agilent 1200 series HPLC system (Agilent Technologies Inc., Santa Clara, CA, USA) with a diode array detector was used.…”

“…Total phenolic content was determined using the modified Folin–Ciocalteu method [ 62 ]. An amount of 0.2 mL of stevia extract was mixed with 1 mL of 0.2 N Folin–Ciocalteu reagent and 0.8 mL 7.5% sodium carbonate solution.…”

“…Total flavonoid content was analyzed by a colorimetric method based on the complexation of phenolic compounds with Al (III) [ 62 ]. An amount of 80 µL of stevia extract was mixed with 1920 µL of a reagent containing 40% ethanol, 0.7% acetic acid, 0.4% hexamethylenetetramine, and 0.6% aluminum chloride.…”

“…Antioxidant activity was measured based on the scavenging of the stable 2,2-diphenyl-1-picrylhydrazyl free radical (DPPH) as described [ 62 ]. Accordingly, 50 µL of stevia extract were mixed with 1950 µL of a DPPH solution (0.025 mg mL, prepared in acetonitrile:methanol:sodium acetate buffer (100 mM, pH 5.5) (1:1:2)).…”