The Stevia rebaudiana plant produces steviol glycosides, a non-caloric sweeteners that are not metabolized in the human body. In addition to the sweet taste, it has various applications for health improvement acts as antihyperglycemic, anticancerous, antihypertensive agent and prevents bacterial and fungi growth [1]. The increasing demand of such products like stevia sweeteners is also putting pressure on the need to improve stevia agronomical traits, such as seed germination, disease resistance and biomass [2]. It has been proven that seed treatment with cold plasma had positive effects on these traits, however changes in the biosynthesis of secondary metabolites are not yet fully understood [3]. The aim of this study was to determine low-pressure plasma (CP)-, dielectric barrier discharge plasma (DBD)-, and vacuum-induced changes in antioxidant activity and concentrations of flavonoids and phenolic compounds in stevia leaves.
The extracts from stevia leaves were prepared by using 70% ethanol and ultrasound (1h) extraction. The concentration of flavonoids was measured using adjusted spectrophotometric FC method [3]. For determination of phenolic compounds another spectrophotometric method was chosen using reagent consisting of 96% ethanol; 33% acetic acid; 5% hexamethylenetetramine and 10% aluminum chloride [3]. DPPH activity measured according to the standardized protocol [3].
Results shows that CP exposure decreases antioxidant activity after 2- and 7-min. exposure (CP2; CP7) compared to unexposed (Control) group, p<0.05. Although longer (CP7) exposure of decreased activity less than shorter exposure length (Table 1). A similar tendency was observed in decrease of phenolic compounds concentration (Table 1). Positive tendency was captioned in flavonoid concentration compared to the control group (Table 1). On the other hand, exposure of DBD shows tendency in flavonoid concentration decrease with longer exposure lengths (Table 1). However, longer DBD exposure (DBD7) increased antioxidant activity (p<0.05). Tendency in phenolic compounds concentration increase after DBD exposure is also consistent with increased antioxidant activity (Table 1).
| Flavonoid conc., mg GAE/g | Phenolic compounds conc., mg GAE/g | Antioxidant activity, mg RUE/g | |
|---|---|---|---|
| Control | 48.8 ± 11.2 | 102.0 ± 18.0 | 124.1 ± 5.0 |
| V5 | 58.8 ± 11.0 | 136.1 ± 4.2 | 111.0 ± 11.3 |
| CP2 | 58.8 ± 6.3 | 55.4 ± 4.9 | 27.5 ± 9.2* |
| CP5 | 61.5 ± 18.7 | 78.0 ± 33.9 | 57.2 ± 30.1 |
| CP7 | 73.7 ± 3.5 | 89.6 ± 13.0 | 69.4±9.7* |
| DBD2 | 86.2 ± 1.7 | 97.3 ± 10.2 | 53.5 ± 21.0 |
| DBD5 | 47.1 ± 4.2 | 62.7 ± 1.8 | 46.3 ± 6.4 |
| DBD7 | 46.0 ± 12.6 | 103.6 ± 19.8 | 142.3 ± 22.5* |
The results suggests that antioxidant activity correlates with the concentration of phenolic compounds rather than the concentration of flavanoids. DBD7 group shows the strongest antioxidant activity, while CP exposure only decreased it. Studies had shown that plasma-induced changes are dependent on the origin of plant and lenght of exposure. It was also confirmed that seeds exposure with CP and DBD has a positive effect on antioxidant activity [4,5,6]. Therefore, the standartization of different plasma application is needed for different plant species. It could be concluded that stevia seeds treatment with dielectric barrier discharge plasma for 7 min has positive effect on antioxidant activity in stevia leaves. However, further research is needed to determine underlying mechanisms of the changes.