HPLC-HRMS/MS Characterization, Molecular Docking, and Comparative Genotoxicity of Free and Liposome-Encapsulated...

Plant extracts remain an important source of antiviral agents, while nanocarrier-based delivery systems have emerged as a promising strategy for improving their therapeutic safety and efficacy. Building on previous findings indicating that nanoliposomal encapsulation improved the anti-coronavirus activity of Aesculus hippocastanum seed extract against human coronavirus strain OC43 (HCoV-OC43), the present study evaluated whether encapsulation reduces the genotoxic effects of the extract and explored a potential molecular basis for the previously observed antiviral activity. Genotoxicity assessment using the alkaline comet assay in MRC-5 human lung fibroblasts demonstrated that at 1 mg/mL, liposomal encapsulation was associated with an approximately 86.0% reduction in %Tail DNA, from 21.301 ± 0.368% for the free extract to 2.984 ± 1.523% for the liposome-encapsulated extract. In parallel, chemical profiling by high-performance liquid chromatography coupled with high-resolution tandem mass spectrometry (HPLC-HRMS/MS) enabled the identification and tentative annotation of 46 metabolites, including triterpenoid saponins, flavonoids, indole-3-acetic acid glycosides, and organic acids. Based on this phytochemical profile, representative constituents were subjected to molecular docking against the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) nonstructural proteins Nsp5 and Nsp12 to investigate their potential contribution to the previously demonstrated antiviral activity of the extract. Among the evaluated compounds, hesperidin exhibited the most favorable predicted binding affinity toward both targets, while quercetin also showed favorable interactions, suggesting that flavonoid constituents may contribute to the antiviral activity of the extract. These findings indicate that liposomal encapsulation improves the biological safety profile of A. hippocastanum seed extract, while phytochemical characterization and molecular docking provide preliminary insights into the constituents that may contribute to its previously demonstrated anti-coronavirus activity.