The aim of this work is to investigate synthesis conditions of water insoluble β-cyclodextrin (β-CD) nanospheres that could be useful as an adsorbent to remove very low concentration of p-nitrophenol (p-NP) from water. The effect of several parameters on β-CD polymer synthesis such as β-CD/epichlorohydrin (EPI) mole ratio, reaction temperature and NaOH concentration was investigated. The FTIR-ATR analysis confirmed the formation of β-CD polymer structure with the addition of EPI by preserving the basic β-CD structural units. To determine the optimal synthesis conditions, performances of the synthesized β-CD nanospheres were determined by measuring the adsorption efficiencies of p-NP. On the basis of these results, the optimal synthesis conditions of β-CD nanospheres were 65°C, 40% NaOH (w/w) concentration and β-CD/EPI=1/55 mole ratio. Langmuir and Freundlich isotherm models were fitted on experimental data. It was found that Langmuir model described best for adsorption of p-NP onto β-CD nanospheres. On the basis of the Langmuir analysis, the maximum adsorption capacities were determined to be 17.203 mg p-NP per gram of β-CD nanospheres. The β-CD nanospheres at predetermined optimum reaction conditions were also characterized by transmission electron microscopy (TEM), scanning electron microscopy (SEM) and Brunauer-Emmet-Teller (BET) analysis. The combination of results from SEM,TEM and BET indicated that the synthesized β-CD nanospheres have a porous and sponge-like structure and 1.54 m2/g specific surface area.