A dual-context assessment of non-target glyphosate responses in Plantago lanceolata L

Urban vegetation can experience chronic, low-intensity glyphosate exposure via management-derived drift within a heterogeneous co-stress background, yet stress-response organisation under such conditions remains insufficiently resolved. We assessed responses of Plantago lanceolata using a dual-context design comprising an urban in situ survey and a standardized ex situ greenhouse exposure, analysed independently. Across three sampling years (2023–2025), we quantified structural traits (histology; stomatal density and size), oxidative–osmoprotective markers including class III peroxidase activity (POD), superoxide dismutase activity (SOD) and proline content, phenolic metabolites and phytohormones by ultra-performance liquid chromatography–tandem mass spectrometry (UPLC–MS/MS), glyphosate and aminomethylphosphonic acid (AMPA) residues by gas chromatography–mass spectrometry (GC–MS), and rhizosphere enzyme activities including acidic phosphatase (ACP), alkaline phosphatase (ALP) and β-glucosidase. In pooled within-context summaries, exposed plants exhibited reduced stomatal density (−42%) and stomatal size (−13%) relative to controls, whereas biomass was lower in exposed plants (10.26 vs. 7.12 g; p < 0.05). Metabolic outputs indicated redistribution within the phenylpropanoid sector (increased p -coumaric acid; decreased vanillin) alongside elevated oxidative defence (POD +102%; SOD +18%) and proline accumulation (+386%). Residues accumulated predominantly in roots, and ACP activity was higher in soils adjacent to exposed plants; however, the in situ survey does not support dose–response inference. Convergent structural and biochemical changes across both contexts support a reproducible, sublethal glyphosate-associated response in P. lanceolata ; however, the in situ survey precludes dose–response inference.