Molecular-LevelInsights into the Interactions ofNicotine and Cotinine with DPPC Phospholipid Bilayers

Abstract Small alkaloids such as nicotine and its major metabolite, cotinine, are widely present in biological and environmental systems, yet their membrane interaction mechanisms and related cytotoxic effects remain poorly quantified. Here, molecular dynamics simulations combined with umbrella sampling were used to investigate the adsorption, distribution, orientation, and transmembrane behavior of nicotine and cotinine in DPPC bilayers. Both molecules rapidly migrated from bulk water to the water–lipid interface within 8 ns and showed strong interfacial enrichment, with residence probabilities of 85% for nicotine and 73% for cotinine. Nicotine exhibited pronounced membrane permeability, repeatedly crossing the entire bilayer, whereas cotinine remained largely confined to the interfacial region. Free energy profiles revealed a favorable minimum for nicotine inside the bilayer (0.89 nm from the center) and a negative transfer free energy (−2 kcal/mol), while cotinine faced a substantial barrier with a positive transfer free energy of 2.5 kcal/mol. Orientational analyses showed broader angular distributions for nicotine, consistent with frequent conformational inversions, whereas cotinine displayed more restricted and stable interfacial orientations. Both molecules reduced lipid acyl-chain order parameters, altered membrane electrostatic potentials, and induced local membrane thinning from an average thickness of ∼3.82 nm. These results quantitatively demonstrate how subtle molecular differences control membrane affinity, permeability, and membrane perturbation, providing fundamental insight into alkaloid–membrane interactions and their potential biological implications.