Molecular networking unveils anti-SARS-CoV-2 constituents from traditionally used remedies

Ethnopharmacological relevance: Plants and fungi have a long tradition in ethnopharmacology for the treatment of infectious diseases including viruses. Many of these natural products have also been used to combat SARS-CoV-2 infections or symptoms of the post- and long-COVID form, owing to the scarcity of clinically approved therapeutics. Aim of the study: The ongoing threat posed by SARS-CoV-2, along with the rapidly evolving new variants, requires the development of new antiviral compounds. The aim of this study was to identify anti-SARS-CoV-2 herbal and fungal extracts used in traditional medicine against acute respiratory infection, inflammation, and related symptoms. Additionally, we sought to characterize their bioactive constituents. Materials and methods: The antiviral activity and cell cytotoxicity of 179 herbal and fungal extracts were evaluated using two SARS-CoV-2 infection assays in Caco-2 cells. 19 plant extracts with and without anti-SARS-CoV-2 activity underwent detailed dereplication using molecular networking. Results: Extracts from Angelica sinensis (Oliv.) Diels roots, Annona squamosa L. seeds, Azadirachta indica A. Juss. fruits, Buddleja officinalis Maxim. flowers, Burkea africana Hook. bark and Clinopodium menthifolium (Host) Stace aerial parts showed a potent anti SARS-CoV-2 activity (IC₅₀ < 5 μg/ml) with only moderate cytotoxicity (CC₅₀ > 60 μg/ml, Caco-2). By performing the dereplication with a bioactivity-featured molecular network (MN) on the extract library level, rather than on the level of individual extracts, we could pinpoint compounds characteristic for active extracts. Thus, a straight-forward identification of potential anti-SARS-CoV-2 natural compounds was achieved prior to any fractionation or isolation efforts. Conclusions: A sophisticated hyphenation of empirical knowledge with MS-based bioinformatics and automated compound annotation was applied to decipher the chemical space of the investigated extracts. The correlation with experimentally assessed anti-SARS-CoV-2 activities helped in predicting compound classes and structural elements relevant for the antiviral activities. Consequently, this accelerated the identification of constituents from the investigated mixtures with inhibitory effects against SARS-CoV-2.