Antifungal efficacy of linoleic-acid-rich Cucurbita pepo L. seed oil revealed by GC-MS profiling of lipids and bioactive markers, ergosterol depletion, and network pharmacology

The development of multi-target, sustainable botanical alternatives is required due to the increasing resistance of phytopathogens to synthetic fungicides. The antifungal effectiveness of Cucurbita pepo L. seed oil (PSO) against a panel of six economically significant phytopathogens is assessed in this study. The PSO was characterized by quantitative GC-MS profiling as a complex matrix dominated by oleic acid (36.52%) and linoleic acid (43.18%), along with diverse bioactive markers such as oxygenated terpenoids and phytosterols. High broad-spectrum activity was shown in vitro, with 100% inhibition of Botrytis fabae and 88.9% inhibition of Fusarium oxysporum at 10 mg·mL⁻¹. Quantitative HPLC analysis confirmed that PSO acts as a sterol-disrupting agent, causing near-complete loss of membrane ergosterol in B. fabae (100% loss) and F. oxysporum (95.2% loss). Furthermore, network pharmacology and molecular docking suggest that the bioactive matrix exerts its effects by destabilizing the fungal cell envelope, with computational models predicting putative interactions with lanosterol 14-alpha-demethylase (CYP51A) and β-1,3-glucan synthase (FKS1). The results position the PSO bioactive matrix as a potent in vitro antifungal agent that targets pathogens directly through membrane destabilization and putative multi-site enzymatic interference. However, detached leaf assays indicate that its future practical application will require formulation strategies, such as nano-emulsions, to mitigate the mild phytotoxicity associated with direct foliar application of the crude oil at fungicidal doses.