A novel flexible pyrene-based chemosensor 1 bearing two benzimidazolium moieties and a urea group was designed and synthesized. The binding properties of chemosensor 1 toward various anions were examined via UV-vis, fluorescence, and ¹H NMR spectroscopic methods. The results demonstrated that chemosensor 1 could distinguish H₂PO₄⁻ from other tested anions such as F⁻, Cl⁻, Br⁻, I⁻, AcO⁻ and HSO₄⁻ by ratiometric fluorescence as chemosensor 1 formed a complex with H₂PO₄⁻ in 2 : 2 ratio via intermolecular excimer transduction in CH₃CN. The fluorescence response of chemosensor 1 to H₂PO₄⁻ was hardly interfered by other anions. The detection limit of chemosensor 1 (5 × 10⁻⁶ M) toward H₂PO₄⁻ is 5.02 × 10⁻⁷ M.