{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,2,27]],"date-time":"2026-02-27T07:25:11Z","timestamp":1772177111145,"version":"3.50.1"},"reference-count":39,"publisher":"MDPI AG","issue":"3","license":[{"start":{"date-parts":[[2026,2,25]],"date-time":"2026-02-25T00:00:00Z","timestamp":1771977600000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/creativecommons.org\/licenses\/by\/4.0\/"}],"funder":[{"DOI":"10.13039\/501100001322","name":"South African Medical Research Council","doi-asserted-by":"crossref","award":["2025\/2026"],"award-info":[{"award-number":["2025\/2026"]}],"id":[{"id":"10.13039\/501100001322","id-type":"DOI","asserted-by":"crossref"}]},{"name":"SAMRC RCDI-Nested PhD Scholarship Programme","award":["2024\/2026"],"award-info":[{"award-number":["2024\/2026"]}]}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Life"],"abstract":"<jats:p>The SARS-CoV-2 papain-like protease (PLpro) is an essential viral enzyme that promotes viral polyprotein processing while simultaneously suppressing the host innate immune response, which makes it a primary target for developing antiviral drugs. The present study employs a comprehensive approach integrating untargeted metabolomic profiling, in silico molecular docking and dynamics simulations, Molecular Mechanics Generalized Born Surface Area (MM-GBSA) energetic assessments, and biochemical enzyme assays. This integrated method aims to discover natural PLpro inhibitors from two ethnomedicinal plants, Lippia javanica and Acorus calamus, which have long been utilized in African traditional medicine to treat respiratory diseases. Comprehensive metabolite profiling using untargeted Ultra-Performance Liquid Chromatography\u2013Tandem Mass Spectrometry (UPLC-MS\/MS) and Global Natural Products Social (GNPS) molecular networking revealed flavonoid glucuronides and phenylpropanoid derivatives as the major constituents in both plant species. In situ histochemical staining further offered spatial validation of phenolic- and lignin-associated tissues, supporting the phenolic-dominated molecular families detected by GNPS molecular networking. In silico evaluation of six selected compounds demonstrated spontaneous and thermodynamically favorable binding to PLpro, with \u0394G_bind values ranging from \u22125.63 to \u22126.43 kcal\/mol. Catechin-7-glucoside emerged as the lead compound, establishing multiple hydrogen bond networks with Asp164, Gln269, Tyr264, and Asn267, supplemented by hydrophobic engagement with Pro247 and Pro248, and \u03c0-\u03c0 stacking with the blocking loop 2 (BL2 loop). Molecular dynamics simulations confirmed the stability of the protein\u2013ligand complexes. Biochemical enzyme assays confirmed concentration-dependent inhibition of PLpro proteolytic and deubiquitinating activity by both crude plant extracts and isolated bioactive compounds. However, S-adenosyl-methionine showed comparatively high PLpro proteolytic activity (IC50 5.872 \u00b5M) compared to catechin-7-glucoside, with an IC50 of 7.493 \u00b5M, exhibiting efficacy similar to the reference inhibitor GRL0617. Both the extracts of L. javanica and A. calamus have shown significant inhibitory activity while maintaining cell viability in Human embryonic kidney 293T cell (HEK293T) culture models, indicating a favorable safety profile of the tested concentrations. Based on these results, catechin-based polyphenols and phenylpropanoid derivatives appear as promising lead compounds for the development of PLpro inhibitors. To progress toward therapeutic use, further work is necessary in pharmacokinetics, structural optimization, and antiviral validation in cell models.<\/jats:p>","DOI":"10.3390\/life16030373","type":"journal-article","created":{"date-parts":[[2026,2,26]],"date-time":"2026-02-26T10:33:02Z","timestamp":1772101982000},"page":"373","update-policy":"https:\/\/doi.org\/10.3390\/mdpi_crossmark_policy","source":"Crossref","is-referenced-by-count":0,"title":["Metabolites from South African Medicinal Plants as Dual-Function Inhibitors of the SARS-CoV-2 Papain-like Protease (PLpro)"],"prefix":"10.3390","volume":"16","author":[{"ORCID":"https:\/\/orcid.org\/0009-0006-6080-1848","authenticated-orcid":false,"given":"Mmamudi Anna","family":"Makhafola","sequence":"first","affiliation":[{"name":"Department of Biology and Environmental Sciences, Sefako Makgatho Health Sciences University, Pretoria 0204, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-9164-8213","authenticated-orcid":false,"given":"Clarissa Marcelle","family":"Naidoo","sequence":"additional","affiliation":[{"name":"Department of Biology and Environmental Sciences, Sefako Makgatho Health Sciences University, Pretoria 0204, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Chikwelu Lawrence","family":"Obi","sequence":"additional","affiliation":[{"name":"Department of Biology and Environmental Sciences, Sefako Makgatho Health Sciences University, Pretoria 0204, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Benson Chuks","family":"Iweriebor","sequence":"additional","affiliation":[{"name":"Department of Biology and Environmental Sciences, Sefako Makgatho Health Sciences University, Pretoria 0204, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-8004-3187","authenticated-orcid":false,"given":"Oyinlola Oluwunmi","family":"Olaokun","sequence":"additional","affiliation":[{"name":"Department of Biology and Environmental Sciences, Sefako Makgatho Health Sciences University, Pretoria 0204, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3048-4304","authenticated-orcid":false,"given":"Earl","family":"Prinsloo","sequence":"additional","affiliation":[{"name":"Department of Biotechnology, Rhodes University, Makhanda 6140, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-6938-0185","authenticated-orcid":false,"given":"Haruhisa","family":"Kikuchi","sequence":"additional","affiliation":[{"name":"Faculty of Pharmacy, Keio University, Tokyo 105-8512, Japan"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Muhammad Sulaiman","family":"Zubair","sequence":"additional","affiliation":[{"name":"Department of Pharmacy, University of Tadulako, Palu 94118, Indonesia"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"ORCID":"https:\/\/orcid.org\/0000-0003-3631-4522","authenticated-orcid":false,"given":"Nqobile Monate","family":"Mkolo","sequence":"additional","affiliation":[{"name":"Department of Biology and Environmental Sciences, Sefako Makgatho Health Sciences University, Pretoria 0204, South Africa"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"1968","published-online":{"date-parts":[[2026,2,25]]},"reference":[{"key":"ref_1","doi-asserted-by":"crossref","first-page":"D608","DOI":"10.1093\/nar\/gkx1089","article-title":"HMDB 4.0: The Human Metabolome Database for 2018","volume":"46","author":"Wishart","year":"2018","journal-title":"Nucleic Acids Res."},{"key":"ref_2","doi-asserted-by":"crossref","first-page":"17","DOI":"10.1186\/1758-2946-4-17","article-title":"Avogadro: An advanced semantic chemical editor, visualization, and analysis platform","volume":"4","author":"Hanwell","year":"2012","journal-title":"J. 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