Fe₃O₄ NPs as novel elicitors for enhanced production of phenylethanoid glycosides in cell suspension culture of Scrophularia striata Boiss

Enhancing the bioactive secondary metabolite production in medicinal plants remains a major challenge, necessitating innovative approaches such as the application of nanoparticles to stimulate metabolic pathways. Among industrially utilized metal nanoparticles, iron oxide nanoparticles (Fe₃O₄ NPs) are recognized as promising novel biostimulants, with applications across diverse fields including medicine, physiology, physics, optics, phytomedicine, food, and health. The potential of Fe₃O₄ NPs to elicit the biosynthesis of valuable phenylethanoid glycosides (PhGs) in Scrophularia striata cell suspension cultures was evaluated. The cells were subjected to different concentrations of Fe₃O₄ NPs (5, 20, 75, and 100 mg/L), and secondary metabolite accumulation was quantified via HPLC. Physiological and biochemical parameters were also evaluated, including cell growth rate, proline content, antioxidant enzyme activities, oxidative stress markers, and total antioxidant capacity. The findings demonstrated that Fe₃O₄ NPs act as potent biostimulants in S. striata cultures, significantly enhancing biosynthesis of secondary metabolites (acteoside, echinacoside, flavonoids, and phenolic acids). These effects are primarily attributed to activation of the shikimate and phenylpropanoid pathways. They are also associated with increased activities of key enzymes, particularly phenylalanine ammonia-lyase and tyrosine ammonia-lyase. The greatest efficacy was found at 5 mg/L, where nanoparticles induced defense-related metabolic responses without causing oxidative damage, suggesting a biphasic dose-response phenomenon, known as hormesis. Conversely, higher concentrations triggered oxidative stress and suppressed biosynthetic pathways. Optimization of nanoparticle concentration is therefore critical to maximize beneficial outcomes while minimizing phytotoxic risks. Results underscore the potential of Fe₃O₄ NPs for enhancing valuable phytochemical production in plant-based biotechnological systems.