Ethephon interactions with biomimetic sperm plasma membranes and their effects on lipid packing and membrane stability.

Ethephon (ETP) is an organophosphorus plant growth regulator extensively used in agriculture, raising concerns about environmental exposure and potential reproductive toxicity. Although animal studies report reduced sperm count, morphological abnormalities, and testicular damage following ETP exposure, the molecular mechanisms underlying these effects remain poorly understood. Because membrane organization and stability are critical determinants of sperm function, we investigated simplified biomimetic sperm plasma membrane systems in the presence of ETP to elucidate membrane-level interaction mechanisms. Lipid assemblies reproducing the main compositional features of sperm membranes (POPC/POPE/sphingomyelin, 2:2:1) were prepared as monolayers and bilayers. Bilayer systems were studied as giant and large unilamellar vesicles (GUVs and LUVs), while interfacial properties were analyzed using Langmuir monolayer isotherms and polarization-modulated infrared reflection-absorption spectroscopy (PM-IRRAS). ETP induced pronounced structural destabilization in GUVs, including possible rupture indicating compromised membrane integrity. In LUVs, LAURDAN fluorescence revealed that ETP significantly modifies the physicochemical environment of the membrane interface, and possibly increases membrane order. At the air-water interface, Langmuir isotherms showed only minor variations in mean molecular area, whereas PM-IRRAS indicated preferential interaction of ETP with lipid polar headgroups accompanied by enhanced vertical ordering of hydrocarbon chains. These results suggest that ETP primarily associates with the membrane interface, where perturbation of headgroup interactions and hydrogen-bonding networks propagates into the bilayer, altering lipid organization and mechanical stability. Overall, the findings demonstrate that ETP directly perturbs membrane physicochemical properties, providing mechanistic insight into how interfacial interactions with lipid assemblies may contribute to impaired sperm membrane function.