Multi-analytical characterization of Stevia rebaudiana powder: nutritional, phytochemical, antioxidant, and NMR-based profiling

The objective of this study was to characterize the Stevia rebaudiana powder in terms of its nutritional composition, phytochemical potential, antioxidant activity, and NMR-based structural profiling. Proximate analysis indicated that Stevia powder contains considerable amounts of protein (10.52 ± 0.21%) and fiber (7.20 ± 0.69%). The mineral profile revealed the presence of Na, K, Mg, Fe, Zn, Ni and Co, with potassium observed as the most abundant mineral. Phytochemical analysis of ethanolic extract indicated considerable quantities of total phenolic (59.48 ± 0.18 mg GAE/g) and total flavonoid (22.91 ± 0.71 mg QE/g) contents, suggesting the presence of bioactive compounds in Stevia rebaudiana powder. The extract exhibited antioxidant activity with DPPH and ABTS inhibition of 80.51 ± 0.96% and 71.22 ± 0.59%, respectively, compared to the standard BHT. Structural analysis of Stevia rebaudiana powder was performed using nuclear magnetic resonance (1H NMR, 13C NMR, and DEPT experiments) spectroscopy. The 1H NMR spectra indicated the signals corresponding to aliphatic and anomeric proton environments, while the 13C NMR spectra provided qualitative information on CH, CH₂, CH₃, olefinic, and carbonyl carbon environments. DEPT experiments showed qualitative differentiation of methine, methylene, and methyl carbon environments within the steviol glycoside framework. Overall, Stevia rebaudiana powder demonstrates notable nutritional composition and antioxidant activity, along with steviol glycoside related signals suggested by NMR analysis. However, spectral overlap and structural complexity limited the definitive identification of individual compounds based only on one-dimensional NMR. This highlights the need for future studies employing orthogonal analytical techniques including quantitative NMR and complementary spectroscopic methods, for detailed characterization of steviol glycosides and their biological relevance.