A new type of magnetic nanoparticles (MNPs), as the absorbents of bisphenol A (BPA), was prepared by functionalization of Fe3O4@SiO2 with BPA-specific aptamer in this work. ssDNA aptamer was immobilized on the Fe3O4@SiO2 surface through biotin-avidin interactions, playing a role of the specific probe for BPA. The resultant materials (Apt-MNPs) exhibited outstanding magnetic responsibility and can be separated efficiently by the magnetic field. Experimental results also showed that Apt-MNPs had large adsorption capacity and high competitive selectivity for the targeted compound BPA. Furthermore, Apt-MNPs were adopted as the specific absorbents to extract and enrich BPA from human serum and urine samples. Therefore, an efficient detection method of BPA was developed in combination with high-performance liquid chromatography (HPLC). The linearity of the method was over a range of 5-10,000 ng mL-1 with a correlation coefficient of 0.99997, and the limit of detections (LODs) for serum and urine were 2.0 and 1.0 ng mL-1, respectively. The recoveries of BPA in the spiked human serum and urine samples were 90.8 ± 7.3% (RSD) and 92.3 ± 1.5%, respectively. Our results demonstrated that Apt-MNPs were high-performance adsorbents for extracting and enriching BPA, resulting in fast and efficient detection of BPA in serum and urine samples. Graphical abstract Aptamer-MNPs were effective for BPA separation from serum and urine.