Penicillium chrysogenum originated chloro-diydropyridyl-oxopropanimidic acid derivative as a potent EPSP synthase-targeted bioherbicide against invasive weed species

Fungi are prolific producers of bioactive metabolites, some of which exhibit herbicidal activity and may serve as sustainable alternatives to synthetic herbicides. In this study, an endophytic Penicillium chrysogenum was investigated for metabolite production. A pure compound, identified as 2-((5-chloro-4-oxo-4, 5-dihydropyridin-3-yl) amino)-3-oxopropanimidic acid (C₈H₈ClN₃O₃; exact mass 229.03), was purified through chromatographic techniques and structurally elucidated using HPLC-MS and detailed 1D/2D NMR analyses. Computational studies revealed strong binding affinity toward EPSP synthase, a validated herbicidal target, with a docking score of - 11.39 kcal/mol. Molecular dynamics and MM-PBSA analyses confirmed the stability of the enzyme-ligand complex, supporting its inhibitory potential. In vitro assays demonstrated significant herbicidal activity against Parthenium hysterophorus and Sonchus oleraceus. At concentrations of 500-1000 µg/mL/mL, the compound completely inhibited seed germination of S. oleraceus and markedly suppressed germination of P. hysterophorus. These results highlight the compound's potential as a bioherbicidal scaffold with dual in silico and in vitro validation. Compared to the synthetic herbicide 2, 4-D, which showed reduced efficacy under identical conditions, this fungal metabolite offers a promising natural and environmentally safer alternative for weed management.