Constructing bivalent aptasensor for sensitive colorimetric detection of amygdalin based on Capture-SELEX screened...

Amygdalin, a cyanogenic glycoside abundant in apricot kernels, poses a food safety risk, requiring rapid detection that conventional chromatographic methods cannot fulfill for on-site application. Thus, this study aims to construct a novel colorimetric sensing platform and improve detection performance through a multivalent aptamer strategy. This pioneering work introduces a novel bivalent aptamer strategy for ultra-sensitive on-site amygdalin detection. The aptamers specific to amygdalin were screened using a five-segment single-stranded DNA library and Capture-SELEX, with truncated variants optimized to obtain a high-affinity core sequence. A key innovation involves the first design and synthesis of a bivalent aptamer that exploits synergistic binding to enhance target affinity. The bivalent aptamer (Bapt 4-4) exhibited doubled affinity for amygdalin compared to its monovalent truncated counterpart (Amyg 4-26), achieving a dissociation constant (K_d) of (2.19 ± 1.03) nM. By integrating the bivalent aptamer with the laccase-like activity of silver oxide nanoparticles (Ag₂O NPs), an aptasensor with high sensitivity and specificity was developed, yielding a detection limit of 0.008 μg mL^-1, which is lower than that of the monovalent aptamer-based sensor (0.0137 μg mL^-1). This multivalent aptamer strategy provides an effective approach for highly sensitive amygdalin detection. Furthermore, the constructed bivalent aptasensor provides a robust technical tool for the on-site monitoring of plant-derived cyanogenic glycoside toxins, with broad application prospects in the domains of food safety and traditional Chinese medicine safety.