Development of non-equilibrium rapid replacement aptamer assay for ultra-fast detection of phthalic acid esters

In this paper, we developed a non-equilibrium rapid replacement aptamer (NERRA) assay that performed ultra-fast (in 30 s) quantitative detection of phthalic acid esters (PAEs) without waiting for the reaction to reach equilibrium. NERRA assay employed fluorescence PoPo3 dye intercalated in an ssDNA aptamer to selectively detect and quantify the PAEs in water. As the intercalated dye was replaced by the PAEs and quenched in the water, the rate of fluorescence change became proportional to PAEs concentration. The sensitivity of NERRA assay was first evaluated with a commercial spectrofluorometer. The selectivity for PAE mixture, individual PAEs, and non-phthalate compounds were also investigated. NERRA assay was also able to quantitatively detect the PAEs in a common plastic product (picnic mat), and the results were compared with those of gas chromatography mass spectrometry. Finally, a custom analyzer (8.5 cm × 8.5 cm × 16.5 cm) was built to demonstrate the portability of the NERRA assay. Using a commercial spectrofluorometer, NERRA assay was able to quantitatively detect a PAE mixture in 30 min with an LOQ of 0.1 μg/L. Using the portable custom analyzer, the detection time was shortened to 30 s with a tradeoff in the LOQ (1 μg/L). In both cases, the LOQs remain within the environmentally relevant PAE concentrations of 0.1-1472 μg/L.