Document Type : Research Paper
Authors
1 Vali e Asr University of Rafsanjan
2 Vali-e-Asr University of Rafsanjan
3 Genetics and Plant Production Dept, Agriculture College, Vali-e-Asr University of Rafsanjan, Rafsanjan, Iran
Abstract
Background and objectives: Urtica dioica L. is a highly valued medicinal plant traditionally used to treat various diseases. Its extracts contain bioactive compounds such as flavonoids, leucoanthocyanidins, and carotene, contributing to its well-documented anticancer, antidiabetic, and antioxidant properties. Given the crucial role of elicitors in enhancing the production of plant secondary metabolites and improving medicinal quality, this study investigates the stimulatory effects of zinc oxide nanoparticles (ZnO-NPs) on the biochemical and metabolic properties of Urtica dioica callus cultures.
Methodology: Nettle (Urtica dioica) seeds were cultured on half-strength Murashige and Skoog (½ MS) medium to obtain sterile seedlings. Internode explants from these seedlings were then excised and transferred to full-strength MS medium supplemented with 1 mg.l-1 benzylaminopurine (BA) and 2.5 mg.l-1 naphthaleneacetic acid (NAA) to induce callus formation. Cultures were maintained under a 16/8 h light/dark photoperiod at 24 °C. Homogeneous calli were obtained through successive subcultures. These calli were then transferred to liquid MS medium and treated with zinc oxide nanoparticles (ZnO-NPs) at 0, 25, and 50 mg.l-1 concentrations. Samples were collected at 12, 24, 48, and 96 hours post-treatment. Biochemical and metabolic parameters, including the activities of catalase (CAT), peroxidase (POD), polyphenol oxidase (PPO), and phenylalanine ammonia lyase (PAL), as well as total protein, total phenolics, quercetin, and kaempferol contents, were measured. The experiment was arranged in a completely randomized design (CRD) using a factorial layout with two factors: ZnO-NP concentration and sampling time, each with three replications.
Results: The results showed that polyphenol oxidase (PPO) activity significantly increased 48 hours after ZnO-NP treatment, particularly at a concentration of 25 mg.l-1. Phenylalanine ammonia lyase (PAL) activity peaked at 12 hours post-treatment at the same concentration (25 mg.l-1). Peroxidase (POD) activity increased only at 12 hours following treatment with 50 mg.l-1 ZnO-NPs, while it declined under all other conditions. Catalase (CAT) activity was highest at 12 hours after ZnO-NP application at 25 mg.l-1. Total protein content increased at two distinct points: 24 hours after treatment with 25 mg.l-1 and 96 hours after treatment with 50 mg.l-1 ZnO-NPs. High-performance liquid chromatography (HPLC) analysis showed no detectable kaempferol at its expected retention time, while quercetin was detected. The highest quercetin accumulation occurred at 25 mg.l-1 ZnO-NPs after 96 hours, and at 50 mg.l-1 after 48 hours of treatment.
Conclusion: This study demonstrated that ZnO-NP elicitation enhanced the activities of key antioxidant and metabolic enzymes, catalase, peroxidase, polyphenol oxidase, and phenylalanine ammonia lyase, as well as increased total protein, total phenolic content, and quercetin accumulation in Urtica dioica callus cultures, depending on concentration and exposure time. These findings suggest that ZnO-NPs can serve as effective elicitors to boost the medicinal and metabolic potential of nettle in vitro. To translate these results into practical applications, further research is needed to optimize large-scale culture systems, such as bioreactors, for cost-effective production.
Keywords
Main Subjects
)