Rhodamine B-assisted fluorescent aptasensor for the sensitive and selective assessment of Doxycycline hyclate in wastewater

Doxycycline (DOX) demonstrates efficacy against SARS-CoV-2; however, its overuse necessitates the development of sensitive and selective sensors due to environmental concerns. Herein, we report a fluorescent aptasensor that uses rhodamine B (RhB), a fluorescent dye that gets quenched upon interaction with DOX. Different parameters like buffer solutions, concentration of Rhodamine B, apt40, and incubation time were optimized and found to be Tris HCl buffer, 4 µM, 25 nM, and 11 min, respectively. Sensor development is facilitated by the aptamer's precise binding to DOX, which quenches the fluorescence emission of dye via hydrogen bond formation between the carboxylic, phenolic, and amino groups of RhB and DOX. The aptasensor demonstrates efficacy across 0.13 µM to 100 µM at pH 7.4, with a detection limit of 114.03 nM. Molecular docking revealed RhB intercalation into ssDNA's minor groove (-3.08 kcal/mol) and multiple hydrogen bonds between DOX and apt40 (-8.43 kcal/mol). The sensor demonstrates excellent selectivity in the presence of interfering agents. Additionally, the real applicability of the sensor was demonstrated by determining DOX in the wastewater sample with 96-100% recovery (RSD < 2%). Its facile preparation, fast analysis, and high sensitivity make this fluorescent aptasensor valuable in environmental monitoring and public health.