We have developed a new competitive protein binding assay (CPBA) based on human serum albumin functionalized silicon dioxide nanoparticles (nano-SiO2-HSA) that can be used for naproxen determination in urine. Compared with a conventional multi-well reaction plate, nano-SiO2 with a high surface-area-to-volume ratio could be introduced as a stationary phase, markedly improving the analytical performance. Nano-SiO2-HSA and horseradish peroxidase-labeled-naproxen (HRP-naproxen) were prepared for the present CPBA method. In this study, a direct competitive binding to nano-SiO2-HSAwas performed between the free naproxen in the sample and HRP-naproxen. Thus, the catalytic color reactions were investigated on an HRP/3,3'5,5'-tetramethylbenzidine (TMB)/H2O2 system by the HRP-naproxen/nano-SiO2-HSA composite for quantitative measurement via an ultraviolet spectrophotometer. A series of validation experiments indicated that our proposed methods can be applied satisfactorily to the determination of naproxen in urine samples. As a proof of principle, the newly developed nano-CPBA method for the quantification of naproxen in urine can be expected to have the advantages of low costs, fast speed, high accuracy, and relatively simple instrument requirements. Our method could be capable of expanding the analytical applications of nanomaterials and of determining other small-molecule compounds from various biological samples.
Keywords: competitive protein binding assay; horseradish peroxidase labeled naproxen; naproxen.