Abstract
A critical challenge in antiviral drug development is identifying virus host interactions while ensuring reproducible clinical data on drug efficacy and safety. These limitations have renewed scientific interest in traditionally used medicinal plants to support host defense, redox balance, and immune regulation. The present study aimed to investigate the antioxidant, SARS-CoV-2 target-modulating activity, pseudovirus entry inhibition, and immunomodulatory effects of Hypericum perforatum extract (HpE) from the Mediterranean region against SARS-CoV-2 and to evaluate its potential as a plant food/nutraceutical bioactive source. The physicochemical properties of HpE were characterized using microtiter assays, HPLC, and LC-MS/MS analysis. To comprehensively determine its cellular effects, we assessed cytotoxicity using the MTT assay, morphological alterations with hematoxylin eosin staining, and examined antioxidant capacity using chemical- and cellular-based assays in Vero E6, HEK293T, Caco-2, and Calu-3 cell lines. To investigate the targeted inhibitory potential of HpE, Receptor Binding Domain (RBD)-Angiotensin Converting Enzyme 2 (ACE2) binding via ELISA and S1 pseudovirus entry in ACE2-overexpressing HEK293T cells were evaluated. Its suppression of SARS-CoV-2 main protease (Mpro) activity was examined using a Fluorescence Resonance Energy Transfer (FRET) assay. Immunomodulatory activity of HpE was investigated by ELISA. HpE exhibited robust antioxidant activity and SARS-CoV-2 relevant inhibitory effects reducing RBD-ACE2 binding by 60% and suppressing Mpro activity by 100% under the tested conditions. Additionally, HpE demonstrated selective and context-dependent immunomodulatory activity, rather than a uniform suppression of inflammatory responses. Overall, these findings indicate that HpE represents a promising plant-derived nutraceutical source for further investigation in SARS-CoV-2 relevant biological models.