Development of an optical probe based on zinc nitride quantum dots for methotrexate determination in biological samples

In this work, a fluorescent nanoprobe has been developed and validated using zinc nitride quantum dots for the sensitive determination of methotrexate (MTX) in human plasma samples. This platform represents the first report of zinc nitride QDs used for MTX sensing, offering an efficient alternative to conventional QDs such as CdTe or ZnO. Given MTX’s narrow therapeutic window and potential toxicity, its accurate monitoring in plasma is clinically essential; however, current analytical methods are often time-consuming and require extensive sample preparation. Zinc nitride QDs were formed by a hydrothermal method involving the reaction of zinc nitrate hexahydrate with ammonia solution. Its morphology and characterization were examined using energy-dispersive X-ray spectroscopy, dynamic light scattering, and transmission electron microscopy. The nanoprobes exhibited a spherical morphology with an average size of < 5 nm. The presence of nitrogen and zinc atoms in the structure of zinc nitride QDs via EDX analysis confirmed the successful synthesis of the nanoprobe. The fluorescence intensity of the developed probe was diminished in the presence of MTX, likely through the electrostatic interactions of carboxyl and amine groups in MTX with the positively charged zinc nitride QDs surface, according to zeta potential measurements. A linear response was observed over the concentration range of 0.02-2.4 µg.mL-1 with a detection limit of 0.003 µg.mL-1. The nanoprobe exhibited relative intra-day and inter-day standard deviations of 1.4 % and 2.3 % (n = 5) for MTX measurements, respectively. Recovery studies in spiked human plasma samples yielded results ranging from 97.0% to 108.0%, confirming the nanoprobe’s accuracy and applicability for real sample analysis. These results indicate the strong potential of zinc nitride QDs as a reliable fluorescent platform for MTX monitoring in clinical settings.