Abstract
Background: A major challenge in antiviral development is the identification of novel virus–host interactions while ensuring therapeutic efficacy and safety. These challenges have renewed interest in phytochemicals derived from medicinal plants as alternative antiviral agents. Objectives: In this study, we investigated the antioxidant, antiviral, and immunomodulatory properties of a Mediterranean Urtica dioica extract (UdE) against SARS-CoV-2 using chemical, biochemical, and in vitro approaches. Methods: The physicochemical properties of UdE were characterized using microtiter assays and HPLC analysis. Cytocompatibility was evaluated in HEK293T, Vero E6, Caco-2, and Calu-3 cell lines while antioxidant activity was assessed using both chemical and cell-based assays. Antiviral activity was evaluated by assessing inhibition of SARS-CoV-2 receptor binding domain (RBD)–ACE2 interaction using ELISA, inhibition of SARS-CoV-2 main protease (Mpro) activity via FRET assay and inhibition of viral entry using SARS-CoV-2 S1 pseudovirus neutralization assay. Results: UdE (100 µg/mL) inhibited RBD–ACE2 binding by 94% and suppressed Mpro activity by 74%, while reducing moderate but significant inhibition of pseudovirus entry (33.6%) at 300 µg/mL dose level in ACE2 expressing HEK293T cells. Immunomodulatory analysis revealed significant suppression of IL-1β and IL-6 production, accompanied by increased TNF-α and IL-8 levels. Conclusions: Collectively, these findings highlight that UdE exhibits multi-target in vitro antioxidant, antiviral, and immunomodulatory activity against SARS-CoV-2; therefore, UdE represents a promising bioactive extract for the management of SARS-CoV-2 infection.