Computational Chemistry-Guided Rational Hapten Design and Elucidation of the Molecular Recognition Mechanism Enable Immunochromatographic Detection of Maltol and Ethyl Maltol in Food and E-Liquid Samples

Abstract Given the potential health risks arising from the abuse or adulteration of maltol (MA) and ethyl maltol (EM)representative synthetic flavor enhancersparticularly in e-liquids, infant formulas, and fraudulently mislabeled foods, there is an urgent need for rapid and sensitive detection methods. Herein, guided by a computational chemistry-based rational hapten design strategy, four haptens were screened, and H4 was identified as the optimal candidate. Using animal immunization combined with hybridoma technology, a high-affinity, dual-specificity monoclonal antibody (mAb-2D6) was generated, exhibiting half-inhibitory concentrations (IC50) of 17.34 ng/mL for MA and 15.97 ng/mL for EM. Molecular docking revealed that key amino acid residuesGly103, His31, Tyr101, Ala104, and Phe99mediate the dual-specific recognition of mAb-2D6 through hydrogen bonds and π-alkyl interactions. Leveraging this mAb-2D6, two immunochromatographic assays (ICAs)a colorimetric format based on gold nanoparticles and a fluorescent format based on quantum dot microsphereswere developed for the first time for on-site rapid detection of MA and EM in soybean oil, infant formula, and e-liquids, with limits of detection ranging from 12.64 to 20.78 μg/kg. The ICA completes the entire detection workflow within 15 min and demonstrates excellent sensitivity, accuracy, practicality, and user-friendliness, enabling deployment in resource-limited field settings.