RESUMO
The growing reliance on pesticides for pest management in agriculture highlights the need for new analytical methods to detect these substances in food and water. Our research introduces a SPRWG-(C18H37) lipopeptide (LP) as a functional analog of acetylcholinesterase (AChE) for glyphosate detection in environmental samples using phosphatidylcholine (PC) monolayers. This LP, containing hydrophilic amino acids linked to an 18-carbon aliphatic chain, alters lipid assembly properties, leading to a more flexible system. Changes included reduced molecular area and peak pressure in Langmuir adsorption isotherms. Small angle X-ray scattering (SAXS) and atomic force microscopy (AFM) analyses provided insights into the LP's structural organization within the membrane and its interaction with glyphosate (PNG). Structural and geometric parameters, as derived from in silico molecular dynamics simulations (MD), substantiated the impact of LP on the monolayer structure and the interaction with PNG. Notably, the presence of the LP and glyphosate increased charge transfer resistance, indicating strong adherence of the monolayer to the indium tin oxide (ITO) surface and effective pesticide interaction. A calibration curve for glyphosate concentration adjustment revealed a detection limit (LOD) of 24 nmol L-1, showcasing the high sensitivity of this electrochemical biosensor. This LOD is significantly lower than that of a similar colorimetric biosensor in aqueous media with a detection limit of approximately 0.3 µmol L-1. Such an improvement in sensitivity likely stems from adding a polar residue to the amino acid chain of the LP.
RESUMO
The large-scale use of glyphosate pesticides in food production has attracted attention due to environmental damage and toxicity risks. Several regulatory authorities have established safe limits or concentrations of these pesticides in water and various food products consumed daily. The irreversible inhibition of acetylcholinesterase (AChE) activity is one of the strategies used for pesticide detection. Herein, we found that lipopeptide sequences can act as biomimetic microenvironments of AChE, showing higher catalytic activities than natural enzymes in an aqueous solution, based on IC50 values. These biomolecules contain in the hydrophilic part the amino acids l-proline (P), l-arginine (R), l-tryptophan (W), and l-glycine (G), covalently linked to a hydrophobic part formed by one or two long aliphatic chains. The obtained materials are referred to as compounds 1 and 2, respectively. According to fluorescence assays, 2 is more hydrophobic than 1. The circular dichroism (CD) data present a significant difference in the molar ellipticity values, likely related to distinct conformations assumed by the proline residue in the lipopeptide supramolecular structure in solution. The morphological aspect was further characterized using small-angle X-ray scattering (SAXS) and cryogenic transmission electron microscopy (cryo-TEM), which showed that compounds 1 and 2 self-assembly into cylindrical and planar core-shell structures, respectively. The mimetic AchE behaviour of lipopeptides was confirmed by Ellman's hydrolysis reaction, where the proline residue in the peptides act as a nucleophilic scavenger of organophosphate pesticides. Moreover, the isothermal titration calorimetry (ITC) experiments revealed that host-guest interactions in both systems were dominated by enthalpically-driven thermodynamics. UV-vis kinetic experiments were performed to assess the inhibition of the lipopeptide catalytic activity and the IC50 values were obtained, and we found that the detection limit correlated with the increase in hydrophobicity of the lipopeptides, implying the micellization process is more favorable.
