Abstract: Nano-biochar (BNPs) commonly coexists with the iron oxide goethite (GT) in aquatic environments, and their hetero-aggregation behavior significantly affects the adsorption of levofloxacin (LEV) on BNPs. As a key water chemistry parameter, solution pH plays a crucial regulatory role in the hetero-aggregation of BNPs and GT, as well as in the migration, transformation, and fate of LEV. To investigate the impact of solution pH on the hetero-aggregation behavior of BNPs and GT in water and its effect on LEV adsorption, various characterization techniques, including X-ray diffraction (XRD), Fourier-transform infrared (FTIR) spectroscopy, and nitrogen adsorption-desorption isotherms, were used to analyze the crystal structure, specific surface area, and surface morphology of the samples. Additionally, aggregation, sedimentation, and adsorption experiments were conducted under different pH conditions to explore the effects of pH on BNPs-GT hetero-aggregation and LEV adsorption. The results revealed that: (1) Under acidic conditions (pH=5-6), BNPs and GT carried opposite surface charges, leading to hetero-aggregation driven by electrostatic attraction. (2) Under alkaline conditions (pH=8-9), the hydrodynamic diameter (D_h) of BNPs-GT hetero-aggregates increased more rapidly, with hydrogen bonding and Lewis acid-base interactions emerging as the primary aggregation mechanisms, indicating that intermolecular forces dominated the hetero-aggregation process. (3) During the adsorption process, solution pH significantly influenced the adsorption performance of the aggregates toward LEV by regulating the surface charge and aggregation behavior of BNPs-GT. The combined effects of hydrogen bonding (both conventional and charge-assisted), electrostatic interactions, and pore filling resulted in the masking or occupation of BNPs′ adsorption sites and functional groups, thereby inhibiting the adsorption of LEV on BNPs. This study demonstrates that solution pH governs the hetero-aggregation mechanism of BNPs and GT by modulating surface charge and intermolecular interactions, while the hetero-aggregation process significantly suppresses BNPs′ adsorption capacity for LEV by masking or occupying adsorption sites and functional groups. These findings provide a theoretical basis for understanding the migration, transformation, and fate of BNPs in aquatic environments, as well as their impact on antibiotic behavior.