Fluoroquinolones (FQs) are veterinary drug residues in the food chain that threaten global food safety and human health and there is a great need for a rapid, sensitive and reliable monitoring method to better safeguard our food systems. In this study, we developed a high-performance aptamer via a recognition-mechanism-oriented truncation strategy, enabling rapid and sensitive detection of multiple FQs. The recognition mechanism between the aptamer and FQs were studied by the combination of computer-aided molecular simulation and experimental verifications. It was revealed that hydrophobicity of FQs critically governs aptamer-target interactions. Guided by these insights, the original sequence was truncated to a 40-nt aptamer, named as W-G1, which preserved key binding sites and structural stability. W-G1 with the characteristic G-quadruplex (G4) conformation demonstrated excellent affinity and stability for FQs. Exploiting the function of this conformation, a label-free fluorescent G4/thioflavine T biosensor was fabricated to detect FQs in chicken ham sausage within 30 minutes. The results showed that detection limits ranged from 1.32 ng/kg to 17.30 ng/kg that are lower than food safety regulation defined limits of 1.0 μg/kg (the lowest LOD of standard method for lomefloxacin), with recoveries of 81.17~115.94%. This mechanism-driven strategy enhances aptamer-based detection of FQs, offering improved performance, design flexibility, and reduced cost.

In this paper, a magnetic separation assisted fluorescence method for the determination of three common organophosphorus pesticides (OPPs) in ‘Shatangju’ mandarin was established. Magnetic beads were synthesized by solvothermal method and modified by polypyrrole (PPy) to form Fe₃O₄@PPy nanoparticles as a fluorescence quenching agent. This method was based on the use of a 6-carboxyfluorescein (FAM)-labeled aptamer (S4-29) by virtue of its broad-spectrum recognition ability for OPPs. When Fe₃O₄/PPy nanoparticles mass ratio, Fe₃O₄@PPy nanoparticles concentration and incubation time were 1:1.5, 0.2 mg/mL, and 120 min, respectivley, the detection limits of phorate, omethoate and profenofos were 4.17 × 10^-4, 2.77 × 10^-4, and 4.67 × 10^-4 μg/kg, and the recoveries ranged from 79.94% to 108.00% with relative standard deviations (RSDs) between 3.05% and 7.39%. In conclusion, this method has strong selectivity and high accuracy and can be used for the analysis of banned OPPs in agricultural products.

Aptamers as a kind of biological recognition element have shown great potential in monitoring and the rapid quantification of organophosphorus pesticides (OPPs). However, molecules of OPPs are structurally similar and original aptamers selected by systematic evolution of ligands by exponential enrichment are usually long-chain bases, which hamper the further application under OPPs-aptamer recognition. The aim of the research was to develop a new strategy to design oligonucleotide sequences for binding OPPs by combination of experimental and molecular modeling methods. 3D models of aptamers binding OPPs were constructed, and binding energy and the most probable binding site for the OPPs were then determined by molecular docking, and the binding sites were further confirmed by the results of 2-AP replaced experiments. Based on the docking results, a new aptamer for detection 4 representative OPPs with only 29 bases was designed by reasonable truncation and mutation of the reported aptamer (named S4-29). The interaction between this new aptamer and OPPs were analyzed by molecular docking, microscale thermophoresis, circular dichroism and fluorometric analysis. The results revealed that the new aptamer exhibit more superior recognition performance to OPPs, which can be promote the monitoring ability of OPPs contaminations in food.